ANNOgesic’s subcommands¶
In general, the subcommands need at least one argument - the analysis folder. If this is not given ANNOgesic assumes that the current folder is the analysis folder.
The format of filename¶
In order to recognize the file types and the relationship between different files, please follow the principle of the format of filename.
Keep the filename of genome fasta file the same as annotation file. For example,
NC_007795.fa, NC_007795.gff, NC_007795.ptt, NC_007795.rnt, NC_007705.gbk
.
All the .gff
files which are import into ANNOgesic
, please follow the format.
$STRAINNAME_$FEATURE.gff
. For example, NC_007795_TSS.gff, NC_007795_transcript.gff
.
The names of features are:
| Feature name | meaning |
| TSS | transcription starting site |
| processing | processing site |
| transcript | transcript assembly |
| sRNA | small RNA |
| sORF | small open reading frame |
| term | terminator |
| 5UTR | 5’UTR |
| 3UTR | 3’UTR |
| circRNA | circular RNA |
| riboswitch | riboswitch |
Please also notice the name of strain or genome in files. It should be avoid to use |
in the name.
Because |
is one character which already used in Unix system. However,
if your fasta files or gff files are download from NCBI through ANNOgesic, ANNOgesic will automatically modify the names.
The input format of libraries for running ANNOgesic¶
Some modules of ANNOgesic
need to import the information of libraries.
For import the information of libraries, please follow this format:
$LIBRARY_FILENAME:$LIBRARY_TYPE:$CONDITION:$REPLICATE:$STRAND
$LIBRARY_FILENAME
means the .wig
file.
$LIBRARY_TYPE
can be tex
(TEX+) or notex
(TEX-) or frag
(fragmented, not for tsspredator
).
$CONDITION
is the index of conditions. Please use 1, 2, 3, ... to represent different conditions.
$REPLICATE
is the index of replicates. Please use a, b, c, ... to represent different replicates.
$STRAND
is the strand of wiggle file. Please use + or -.
For example,
for TEX +/- treated libraries:
TSB_OD_0.2_TEX_reverse.wig:tex:1:a:-,
TSB_OD_0.5_TEX_reverse.wig:tex:2:a:-,
TSB_OD_0.2_TEX_forward.wig:tex:1:a:+,
TSB_OD_0.5_TEX_forward.wig:tex:2:a:+,
TSB_OD_0.2_reverse.wig:notex:1:a:-,
TSB_OD_0.5_reverse.wig:notex:2:a:-,
TSB_OD_0.2_forward.wig:notex:1:a:+,
TSB_OD_0.5_forward.wig:notex:2:a:+,
All the libraries should be separated by comma like the above. The libraries should be combined to one line.
for fragmented libraries:
fragmented_forward.wig:frag:1:a:+,fragmented_reverse.wig:frag:1:a:-
The format of sRNA database¶
If you want to import sRNA database to sRNA detection, please follow the format. Or the output of sRNA name will become nonsense. The format is
- ::
- $ID|$STRAIN|$SRNANAME
for example
- ::
- srn_4840|S._aureus_NCTC8325|RsaOV
Definition of reference strain and target strain¶
We call the strain that user want to compute - “target strain”. If user have no fasta file or genome annotation files of “target strain”, ANNOgesic can generate them. It requires a strain which is close to “target strain”. We call this required strain - “reference strain”.
Riboswitch dataset of Rfam¶
For riboswitch detection, it need the information of riboswitch of Rfam. The input format is like the following.
All columns are splited by tab
. You can also download the data from our
Github.
create¶
create
generates the required folders for input and
output files. Once these folders are created, the input files have to
be placed into the correct locations. Therefore, after creating the folders,
please put the required files in the proper folders.
BAMs: For .bam
files. It has two sub-folders. BAMs_map_reference
is for the .bam
files which mapped on “reference strain”.
BAMs_map_target
is for the .bam
files which mapped on “target strain”.
database: For all databases
manual_TSS: If you detected transcription starting sites(TSSs) manually,
you can put the results here. When you compute TSSs, ANNOgesic
can merge manual and predicted TSSs together.
If you want to run TSS_optimization
for TSSs, it is also necessary.
Please use gff3 format.
manual_processing_site: It is similar with manual_TSS
, it is for
processing sites.
mutation_table: If you detected the mutations between reference genome and
target genome manually, please put the file here. When
you run get_target_fasta
, it is required. Please refer
to the section of get_target_fasta
for the format of
mutation table.
reads: If you want to run circrna
and you also want to align read data by ANNOgesic,
please put the read data here. It can also deal with .bzip2
and .gzip
.
reference: For annotation files and fasta files. If the information of
the reference strain can be download from NCBI, you can also get
the files through running get_input_files
.
riboswitch_ID: For storing the file which contains all the Rfam ID of riboswitch.
For the details of format, please refer to the section of
riboswicth
.
wigs: For wiggle files. Based on the methods of RNA-Seq, you can put them to
fragment
(fragmented libraries) or tex_notex
(TEX +/- treated libraries).
- Arguments
usage: ANNOgesic.py create [-h] project_path
positional arguments:
project_path Name/path of the project.
optional arguments:
-h, --help show this help message and exit
get_input_files¶
get_input_files
is the subcommand for downloading required files (fasta, annotation files) from NCBI.
Therefore, user needs to assign the IP of the reference genome in NCBI. For example,
ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCF_000013425.1_ASM1342v1
Then, user can choose which kinds of files he/she wants to download.
- Pre-reqired information
FTP source
: The IP of NCBI.
- Arguments
usage: annogesic get_input_files [-h] [--FTP_path FTP_PATH] [--ref_fasta]
[--ref_gff] [--ref_ptt] [--ref_rnt]
[--ref_gbk] [--convert_embl] [--for_target]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--FTP_path FTP_PATH, -F FTP_PATH
Path of website where can download the required files.
--ref_fasta, -f Download fasta files of reference. Default is False.
--ref_gff, -g Download gff files of reference. Default is False.
--ref_ptt, -p Download ptt files of reference. Default is False.
--ref_rnt, -r Download rnt files of reference. Default is False.
--ref_gbk, -k Download genbank files of reference. Default is False.
--convert_embl, -e Convert gbk to embl files of reference. Default is
False.
--for_target, -t If the genome which you download from NCBI is your
query sequence(you won't modify the genome), you can
assign the download files to store in target folder in
stead of reference folder.
- Output files
The output files will store in $ANNOgesic_folder/input/reference
if --for_target
is False.
The output files will store in $ANNOgesic_folder/output/target
if --for_target
is True.
fasta
: Fasta files.
annotation
: Annotation files.
get_target_fasta¶
get_target_fasta
is the subcommand for generating fasta files of “target strain” from
“reference strain”. The example of format of mutation table is following:
| #target_id | reference_id | reference_nt | position | target_nt | impact of correction | locus tag | gene | Description |
| HG003 | NC_007795.1 | a | 333 | c | SAOUHSC_00002 | dnaA | XXXXXX | |
| HG003 | NC_007795.1 | t | 543 | - | deletion | YYYYYY | ||
| HG003 | NC_007795.1 | - | 600 | g | insertion | SAOUHSC_00132 |
If user wants to put the titles of columns in the top, it needs to start from #
.
Each column is separated by tab
. If the mutation type is deletion or insertion,
user can put -
to represent them. The information of target_id
, reference_id
,
reference_nt
, position
, target_nt
is required. The others can be blank.
However, please still use tab
to separate all blank columns.
If user has no mutation information, user can also use SNP_calling
(one module of ANNOgesic
) to compute it. Please refer to the section of SNP_calling
.
- Pre-required files
Fasta files of reference genome.
Mutation table which indicates the information of mutations between reference and target strain.
- Arguments
usage: annogesic get_target_fasta [-h] [--ref_fasta_folder REF_FASTA_FOLDER]
[--mutation_table MUTATION_TABLE]
[--output_format OUTPUT_FORMAT]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--ref_fasta_folder REF_FASTA_FOLDER, -r REF_FASTA_FOLDER
The path of the folder of fasta files.
--mutation_table MUTATION_TABLE, -m MUTATION_TABLE
The path of mutation table.
--output_format OUTPUT_FORMAT, -o OUTPUT_FORMAT
Please assign the output filename and which strain
should be included in it. For example:
FILE1:strain1_and_strain2,FILE2:strain3. FILE1 is a
output fasta file which include the information of
strain1 and strain2 (import multi-strains to one file
should be separated by "_and_".) And FILE2 is for
strain3. Comma is for split the files.
- Output files
Fasta files of target genome will store in $ANNOgesic_folder/output/target/fasta
.
annotation_transfer¶
annotation transfer
is the subcommand for transfering the annotation from “reference strain”
to target “strain”. In this subcommand, we use PAGIT and RATT
to achieve it. The similarity of “reference strain” and “target strain” should be closed.
Or it will influence the final results.
Please be attation, before you start to run RATT(annotation transfer),
run source $PAGIT_HOME/sourceme.pagit
first. it will modify the path for execute RATT.
If you use Docker to execute ANNOgesic, you can skip the path modification.
- Pre-required tools and files
The genbank files of reference genome.
The fasta files of reference genome.
The fasta files of target genome.
- Arguments
usage: annogesic annotation_transfer [-h] [--RATT_path RATT_PATH]
[--compare_pair COMPARE_PAIR]
[--element ELEMENT]
[--transfer_type TRANSFER_TYPE]
[--ref_embl_gbk REF_EMBL_GBK]
[--ref_fasta REF_FASTA]
[--target_fasta TARGET_FASTA]
[--convert_to_gff_rnt_ptt]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--RATT_path RATT_PATH
Path of the start.ratt.sh file of RATT folder. Default
is start.ratt.sh.
--compare_pair COMPARE_PAIR, -p COMPARE_PAIR
Please assign the name of strain pairs. ex.
NC_007795:NEW_NC_007795. The reference strain is
NC_007795 and the target strain is NEW_NC_007795.
Please assign the names of strain, not filenames of
fasta files. If you want to assign multiple strains,
please use comma to separate the strains.
--element ELEMENT, -e ELEMENT
It will become the prefix of all output file.
--transfer_type TRANSFER_TYPE, -t TRANSFER_TYPE
The transfer type for running RATT.(details can refer
to the manual of RATT.) Default is Strain.
--ref_embl_gbk REF_EMBL_GBK, -re REF_EMBL_GBK
The folder which stores every reference embl
folders.If you have no embl folder, you can assign the
folder of genbank.
--ref_fasta REF_FASTA, -rf REF_FASTA
The folder of reference fasta files.
--target_fasta TARGET_FASTA, -tf TARGET_FASTA
The folder which stores target fasta files.
--convert_to_gff_rnt_ptt, -g
Do you want to convert to gff, rnt and ptt? Default is
False.
- Output files
All the output files from PAGIT and RATT
will store in $ANNOgesic_folder/output/annotation_transfer
.
All annotation files(.gff
, .ptt
, .rnt
) will store in $ANNOgesic_folder/output/target/annotation
.
snp¶
snp
can detect the potential mutations by comparing the results of alignment and fasta files.
Samtools, Bcftools are the main tools
for detection of mutations. User can choose programs (with BAQ, without BAQ and extend BAQ) to run snp
.
User can also set the quality, read depth and the fraction of maximum read depth which support for indel.
If you have no fasta file of “target strain” and want to generate it, snp
can get it from
the alignment results of “reference strain”.
- Pre-required files and tools:
BAM files for fragmented libraries or TEX +/- treated libraries.
Reference or target genome fasta files.
- Arguments
usage: annogesic snp [-h] [--samtools_path SAMTOOLS_PATH]
[--bcftools_path BCFTOOLS_PATH] [--bam_type BAM_TYPE]
[--program PROGRAM] [--fasta_path FASTA_PATH]
[--tex_bam_path TEX_BAM_PATH]
[--frag_bam_path FRAG_BAM_PATH] [--quality QUALITY]
[--read_depth READ_DEPTH]
[--indel_fraction INDEL_FRACTION] [--ploidy PLOIDY]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--samtools_path SAMTOOLS_PATH
If you want to assign the path of samtools, please
assign here.
--bcftools_path BCFTOOLS_PATH
If you want to assign the path of bcftools, please
assign here.
--bam_type BAM_TYPE, -t BAM_TYPE
Please assign the type of BAM. If your BAM file are
mapped to reference strain and you want to know the
mutations between refenece strain and target strain,
plase keyin 'reference'. If your BAM file are mapped
to target strain and you want to check the mutations
of genome sequence, please keyin 'target'.
--program PROGRAM, -p PROGRAM
Please assign the program for detecting SNP of
transcript: 1: calculate with BAQ, 2: calculate
without BAQ, 3: calculate with extend BAQ. You can
assign more than 1 program. For example: 1,2,3.
Default is 1,2,3.
--fasta_path FASTA_PATH, -f FASTA_PATH
The path of genome fasta folder.
--tex_bam_path TEX_BAM_PATH, -tw TEX_BAM_PATH
The path of tex+/- wig folder.
--frag_bam_path FRAG_BAM_PATH, -fw FRAG_BAM_PATH
The path of fragmented wig folder.
--quality QUALITY, -q QUALITY
The minimum quality which considers a real snp.
Default is 40.
--read_depth READ_DEPTH, -d READ_DEPTH
The minimum read depth. Default is 100.
--indel_fraction INDEL_FRACTION, -imf INDEL_FRACTION
The fraction of maximum read depth, which supports
insertion of deletion. Default is 0.5.
--ploidy PLOIDY, -pl PLOIDY
haploid or diploid. Default is haploid.
- Output files
If bam_type
is reference
,
the results will store in $ANNOgesic/output/SNP_calling/compare_reference
.
If it is target
, the results will store in $ANNOgesic/output/SNP_calling/validate_target
.
The raw data from Samtools and Bcftools
will store in $ANNOgesic/output/SNP_calling/$BAM_TYPE/SNP_raw_outputs
.
The results will store in $ANNOgesic/output/SNP_calling/$BAM_TYPE/SNP_table
.
The meaning of file names are:
$STRAIN_$PROGRAM_depth_only.vcf
means the results only match the condition of read depth.
$STRAIN_$PROGRAM_depth_quality.vcf
means the results match the condition of read depth and quality.
$STRAIN_$PROGRAM_seq_reference.csv
is the index of fasta files which generated by snp
.
For example,
Staphylococcus_aureus_HG003 1632629 . AaA AA 57 .
Staphylococcus_aureus_HG003 1632630 . aA a 57 .
Staphylococcus_aureus_HG003 1499572 . T TT,TTTTT 43.8525 .
These mutations will cause conflict. Then the conflict will effect the positions of other mutations.
Therefore, it will generate four different fasta files.
$STRAIN_$PROGRAM_seq_reference.csv
is the index for these four fasta files.
1 1632629 1 1499572:TT Staphylococcus_aureus_HG003
1 1632629 2 1499572:TTTTT Staphylococcus_aureus_HG003
2 1632630 1 1499572:TT Staphylococcus_aureus_HG003
2 1632630 2 1499572:TTTTT Staphylococcus_aureus_HG003
The first column is the index of conflict. The second column is the positions which be selected.
The third one is the index of two potential mutations in the same position. The fourth one is
the position and nucleotides of mutations. The last column is the name of strain.
If you refer to $ANNOgesic/output/SNP_calling/$BAM_TYPE/seqs
, the filename of fasta is like
$FILENAME_$STRIANNAME_$INDEXofCONFLICT_$INDEXofTWOMUTATION.fa
. Therefore, the first line of
$STRAIN_$PROGRAM_seq_reference.csv
will generate
Staphylococcus_aureus_HG003_Staphylococcus_aureus_HG003_1_1.fa
(if the file name of genome is Staphylococcus_aureus_HG003). The second line will generate
Staphylococcus_aureus_HG003_Staphylococcus_aureus_HG003_1_2.fa
and so forth.
The statistics files will store in $ANNOgesic/output/SNP_calling/$BAM_TYPE/statistics
.
tsspredator(TSS and processing site prediction)¶
tsspredator
can generate the candidates of TSSs and processing sites. The main tool is
TSSpredator. We can easily switch the
TEX+ libraries and TEX- libraries to detect processing sites. User can assign the parameters
of TSSpredator. If User want to get the
optimized parameters of TSSpredator,
there is optimize_tsspredator
for this purpose. Please refer to the section of optimize_tsspredator
.
For the information of libraries, please refer to the section
The format of libraries for import to ANNOgesic
.
- Pre-required tools and files
The libraries and wiggle files of TEX +/-. Please refer to The format of libraries for import to ANNOgesic
.
Fasta file of genome sequence.
Gff file of genome annotation.
If user has gff file of manual detected TSSs, tsspredator
can merge the manual one
and predicted one.
If user wants to compare TSSs with transcripts, it also need the gff files of transcripts.
For the transcripts, please refer to the section of transcript_assembly
.
- Arguments
usage: annogesic tsspredator [-h] [--TSSpredator_path TSSPREDATOR_PATH]
[--fasta_folder FASTA_FOLDER]
[--annotation_folder ANNOTATION_FOLDER]
[--wig_folder WIG_FOLDER] [--height HEIGHT]
[--height_reduction HEIGHT_REDUCTION]
[--factor FACTOR]
[--factor_reduction FACTOR_REDUCTION]
[--enrichment_factor ENRICHMENT_FACTOR]
[--processing_factor PROCESSING_FACTOR]
[--base_height BASE_HEIGHT]
[--replicate_match REPLICATE_MATCH]
[--utr_length UTR_LENGTH] [--lib LIB]
[--output_prefix OUTPUT_PREFIX]
[--merge_manual MERGE_MANUAL] [--statistics]
[--validate_gene]
[--compute_program COMPUTE_PROGRAM]
[--compare_transcript_assembly COMPARE_TRANSCRIPT_ASSEMBLY]
[--fuzzy FUZZY] [--cluster CLUSTER]
[--length LENGTH] [--re_check_orphan]
[--overlap_feature OVERLAP_FEATURE]
[--reference_gff_folder REFERENCE_GFF_FOLDER]
[--remove_low_expression REMOVE_LOW_EXPRESSION]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--TSSpredator_path TSSPREDATOR_PATH
If you want to assign the path of TSSpredator, please
assign here. Default is /usr/local/bin/TSSpredator.jar
--fasta_folder FASTA_FOLDER, -f FASTA_FOLDER
Path of the target genome fasta folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
Path of the target genome annotation gff folder.
--wig_folder WIG_FOLDER, -w WIG_FOLDER
The folder of the wig folder.
--height HEIGHT, -he HEIGHT
This value relates to the minimal number of read
starts at a certain genomic position to be considered
as a TSS candidate. Default is 0.3.
--height_reduction HEIGHT_REDUCTION, -rh HEIGHT_REDUCTION
When comparing different strains/conditions and the
step height threshold is reached in at least one
strain/condition, the threshold is reduced for the
other strains/conditions by the value set here. This
value must be smaller than the step height threshold.
Default is 0.2.
--factor FACTOR, -fa FACTOR
This is the minimal factor by which the TSS height has
to exceed the local expression background. Default is
2.0.
--factor_reduction FACTOR_REDUCTION, -rf FACTOR_REDUCTION
When comparing different strains/conditions and the
step factor threshold is reached in at least one
strain/condition, the threshold is reduced for the
other strains/conditions by the value set here. This
value must be smaller than the step factor threshold.
Default is 0.5.
--enrichment_factor ENRICHMENT_FACTOR, -ef ENRICHMENT_FACTOR
This is the minimal enrichment factor. During
optimization will never larger than this value.
Default is 2.0.
--processing_factor PROCESSING_FACTOR, -pf PROCESSING_FACTOR
This is the minimal processing factor. If untreated
library is higher than the treated library and above
which the TSS candidate is considered as a processing
site and not annotated as detected. During
optimization will never larger than this value.
Default is 1.5.
--base_height BASE_HEIGHT, -bh BASE_HEIGHT
This is the minimal number of reads should be mapped
on TSS. Default is 0.0.
--replicate_match REPLICATE_MATCH, -rm REPLICATE_MATCH
The TSS candidates should match to how many number of
the replicates. Default is 1.
--utr_length UTR_LENGTH, -u UTR_LENGTH
The length of UTR. It is for Primary and Secondary
definition. Default is 300.
--lib LIB, -l LIB The libraries of wig files for TSSpredator. The format
is: wig_file_name:tex_treat_or_not(tex or notex):condi
tion_id(integer):replicate_id(alphabet):strand(+ or
-). If you have multiple wig files, please use comma
to separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--output_prefix OUTPUT_PREFIX, -p OUTPUT_PREFIX
The output prefix of all conditions. If you have
multiple conditions, please use comma to separate
them. For example,
prefix_condition1,prefix_condition2.
--merge_manual MERGE_MANUAL, -m MERGE_MANUAL
If you have gff file of manual checked TSS, you can
use this function to merge manual checked ones and
predicted ones. please assign the path of gff file
of manual checked TSS.
--statistics, -s Doing statistics for TSS candidates. it will store in
statistics folder. Default is False.
--validate_gene, -v Using TSS candidates to validate genes in annotation
file. it will store in statistics folder. Default is
False.
--compute_program COMPUTE_PROGRAM, -t COMPUTE_PROGRAM
Which program do you want to predict. (TSS or
processing_site) Default is TSS.
--compare_transcript_assembly COMPARE_TRANSCRIPT_ASSEMBLY, -ta COMPARE_TRANSCRIPT_ASSEMBLY
If you want to compare with transcriptome assembly,
please assign the folder of gff file of transcript
assembly. Default is False.
--fuzzy FUZZY, -fu FUZZY
The fuzzy for comparing TSS and transcript assembly.
Default is 5.
--cluster CLUSTER, -c CLUSTER
This number is for compare manual detected TSS and
prediced one. If the position between manual checked
one and predicted one is smaller or equal than this
value, It will only print one of them. Default is 2.
--length LENGTH, -le LENGTH
The length of genome that you want to compare between
predicted one and manual checked one for statistics.
If you want to compare whole genome, please don't turn
it on. The default is comparing whole genome.
--re_check_orphan, -ro
If your annotation file lack information of gene or
locus_tag, you can turn it on. It will try to compare
with CDS. Default is False.
--overlap_feature OVERLAP_FEATURE, -of OVERLAP_FEATURE
If processing site and TSS are overlap, you can keep
"TSS" or "processing_site" or "both". Default is both.
--reference_gff_folder REFERENCE_GFF_FOLDER, -rg REFERENCE_GFF_FOLDER
For --overlap_feature, if you want to only keep "TSS"
or "processing_site", you need to assign the
--reference_gff_folder. If you are running TSS, please
assign the folder of processing site. If you are
running processing_site, please assign the folder of
TSS. If you want to keep "both" at overlap position,
please don't turn it on. Default is None(for keep
both).
--remove_low_expression REMOVE_LOW_EXPRESSION, -rl REMOVE_LOW_EXPRESSION
If you want to remove low expressed TSS/processing
site, please assign the file of manual checked gff
file here. It will remove the low expressed ones
based on comparisone of manual checked ones.
Please Be ATTENTION: this parameter may
remove some True positive, too. So, please make sure
you want to do it.
- Output files
The output files will be stored in $ANNOgesic/output/TSS
.
MasterTables
: The MasterTable from TSSpredator.
statistics
: Statistics files.
The output files of processing sites are similar. Just replace TSS
to processing_site
like $ANNOgesic/output/processing_site
.
configs
: The configuration files for running TSSpredator.
gffs
: The gff files of TSSs.
There are some useful tags in the attributes of gff files:
Method
: The TSSs are from manual detection or TSSpredator.
type
: The type of TSSs. It could be Primary, Secondary, Internal, Antisense or Orphan.
UTR_length
: The length of UTR.
associated_gene
: Which genes are associated with this TSS.
Parent_tran
: Which transcript are associated with this TSS, if user has compared with transcript.
If user has compared with genome annotation files, the tag - start_TSS
will appear in the gff files
of genome annotation. It represents the TSSs which associates with the CDS/tRNA/rRNA.
If user has compared with transcripts, the tag - associated_tss
will appear in the gff files
of transcript. It will show the associated TSSs which is in the transcript.
transcript_assembly¶
transcript_assembly
will detect transcripts based on the coverage.
For importing the information of libraries, please refer to the section of
The format of libraries for import to ANNOgesic
.
- Pre-required tools and files
Wiggle files of fragmented libraries or TEX +/- treated libraries. We don’t recommend that only use TEX +/- treated libraries to generate transcripts. It will lose some information in 3’end.
If user wants to compare transcripts with TSSs, it requires .gff
files of TSSs.
If user wants to compare transcripts with genome anntation, it requires .gff
files of genomes.
- Arguments
usage: annogesic transcript_assembly [-h]
[--annotation_folder ANNOTATION_FOLDER]
[--length LENGTH]
[--tex_wig_path TEX_WIG_PATH]
[--frag_wig_path FRAG_WIG_PATH]
[--height HEIGHT] [--width WIDTH]
[--tolerance TOLERANCE]
[--tolerance_coverage TOLERANCE_COVERAGE]
[--replicates_tex REPLICATES_TEX]
[--replicates_frag REPLICATES_FRAG]
[--tex_notex TEX_NOTEX]
[--compare_TSS COMPARE_TSS]
[--compare_genome_annotation COMPARE_GENOME_ANNOTATION]
[--compare_feature_genome COMPARE_FEATURE_GENOME]
[--TSS_fuzzy TSS_FUZZY]
[--Tex_treated_libs TEX_TREATED_LIBS]
[--fragmented_libs FRAGMENTED_LIBS]
[--table_best]
[--terminator_folder TERMINATOR_FOLDER]
[--fuzzy_term FUZZY_TERM]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
It is for comparing transcript assembly and genome
annotation gff file. It can use annotation gff file
as reference and modify transcript assembly file.
If you want to do it, please assign the annotation
gff folder. Otherwise, don't turn it on.
--length LENGTH, -l LENGTH
The minimum width of transcript. It is for comparing
to annotation file(--annotation_folder). If you want
to compare with annotation files, it will be the final
output. If you don't want to compare with annotation
files, --width would be minimum length for the final
output. The default is 20.
--tex_wig_path TEX_WIG_PATH, -tw TEX_WIG_PATH
The path of TEX+/- wig folder.
--frag_wig_path FRAG_WIG_PATH, -fw FRAG_WIG_PATH
The path of fragment wig folder.
--height HEIGHT, -he HEIGHT
The minimum height of coverage to be a transcript. The
default is 10.
--width WIDTH, -w WIDTH
The minimum width of transcript. It is for not
comparing to annotation file(--annotation_folder). If
you don't want to compare with annotation files, it
will be the final output. Otherwise, --length would be
the minimum length of transcript for the final output.
The default is 20.
--tolerance TOLERANCE, -t TOLERANCE
This number indicates how many nucleotides which
coverages drop below --height can be ignore. The
default is 5.
--tolerance_coverage TOLERANCE_COVERAGE, -tc TOLERANCE_COVERAGE
If the coverage is lower than tolerance_coverage, even
the range is within --tolerance, it will terminate the
current transcript. The default is 0.
--replicates_tex REPLICATES_TEX, -rt REPLICATES_TEX
The position is included in the current transcript if
the supported replicates are more than --replicates_tex.
(for tex +/- library)
--replicates_frag REPLICATES_FRAG, -rf REPLICATES_FRAG
The position is included in the current transcript if
the supported replicates are more than --replicates_frag.
(for fragmented library)
--tex_notex TEX_NOTEX, -te TEX_NOTEX
If you use tex +/- libraries to run transcript
assembly, the transcripts should be detected by both
or just one. (1 or 2). Default is 1.
--compare_TSS COMPARE_TSS, -ct COMPARE_TSS
If you want to compare with TSS, please assign TSS
folder.
--compare_genome_annotation COMPARE_GENOME_ANNOTATION, -cg COMPARE_GENOME_ANNOTATION
If you want to compare with genome annotation file,
please assign genome annotation folder.
--compare_feature_genome COMPARE_FEATURE_GENOME, -cf COMPARE_FEATURE_GENOME
If you want to compare with genome annotation file,
please assign the feature which you want to compare.
Default is gene. If you want to compare more than one
feature, just insert comma between each feature,
such as gene,CDS.
--TSS_fuzzy TSS_FUZZY, -fu TSS_FUZZY
The fuzzy for comparing TSS and transcript assembly.
Default is 5.
--Tex_treated_libs TEX_TREATED_LIBS, -tl TEX_TREATED_LIBS
Tex +/- library. The format is:
wig_file_name:tex_treat_or_not(tex or notex):condition
_id(integer):replicate_id(alphabet):strand(+ or -). If
you have multiple wig files, please use comma to
separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--fragmented_libs FRAGMENTED_LIBS, -fl FRAGMENTED_LIBS
Fragmented library. The format is: wig_file_name:fragm
ented(frag):condition_id(integer):replicate_id(alphabe
t):strand(+ or -). If you have multiple wig files,
please use comma to separate the wig files. For
example, wig1:frag:1:a:+,wig2:frag:1:a:-.
--table_best, -tb The output table only includes the best library.
Default is False.
--terminator_folder TERMINATOR_FOLDER, -tr TERMINATOR_FOLDER
If you want to compare between transcripts and
terminators, you can assign the folder of gff files of
terminator here. Default is None.
--fuzzy_term FUZZY_TERM, -fz FUZZY_TERM
If you want to compare between transcripts and
terminators, please assign the fuzzy here. Default is
30.
- Output files
The output files will be stored in $ANNOgesic/output/transcriptome_assembly
.
gffs
: The gff files of transcript.
tables
: For more details of transcripts.
statistics
: Statistics files.
There are some useful tags in gff files.
compare_FEATURE
: The situation of overlap between transcripts and features(–compare_feature_genome)
(If user has compared transcripts with genome annotation.)
associated_tss
: Which TSSs are located in this transcripts.
(If user has compared transcripts with TSSs.)
associated_$FEATURE
: It shows the feature(–compare_feature_genome) which are located in this transcripts.
(If user has compared transcripts with genome annotations.)
detect_lib
: The transcript is detected by tex-treated libraries or fragmented libraries.
best_avg_coverage
: The average coverage of highest expressed library.
If user has compared transcripts with genome annotations. The tag - Parent_tran
will appear
in the gff files of genome annotations. It will show which transcirpt that CDSs/tRNAs/rRNAs are located.
If user has compared transcripts with TSSs. The tag - Parent_tran
will appear
in the gff files of TSSs. It will show which transcripts that TSSs are located.
terminator¶
terminator
will predict the rho-independent terminators. ANNOgesic
combine the results of
two methods in order to get more reliable candidates. First one is using TranstermHP.
The other one is detect the specific secondary structure between converging pairs
of transcripts and CDSs. ANNOgesic
can also compare with coverages in order to generate the terminators
which has coverage significant decreasing.
- Pre-required tools and files
RNAfold of ViennaRNA.
Gff files target genome annotation.
Fasta files of target genome sequence.
Wiggle files of TEX +/- treated libraries or fragmented libraries. we don’t recommand only use TEX +/- treated libraries. Because it will lose some information in 3’end.
Gff files of transcript.
- Arguments
usage: annogesic terminator [-h] [--TransTermHP_path TRANSTERMHP_PATH]
[--expterm_path EXPTERM_PATH]
[--RNAfold_path RNAFOLD_PATH]
[--fasta_folder FASTA_FOLDER]
[--annotation_folder ANNOTATION_FOLDER]
[--transcript_folder TRANSCRIPT_FOLDER]
[--sRNA SRNA] [--statistics]
[--tex_wig_folder TEX_WIG_FOLDER]
[--frag_wig_folder FRAG_WIG_FOLDER]
[--decrease DECREASE]
[--fuzzy_detect_coverage FUZZY_DETECT_COVERAGE]
[--fuzzy_within_transcript FUZZY_WITHIN_TRANSCRIPT]
[--fuzzy_downstream_transcript FUZZY_DOWNSTREAM_TRANSCRIPT]
[--fuzzy_within_gene FUZZY_WITHIN_GENE]
[--fuzzy_downstream_gene FUZZY_DOWNSTREAM_GENE]
[--highest_coverage HIGHEST_COVERAGE]
[-tl TEX_NOTEX_LIBS] [-fl FRAG_LIBS]
[-te TEX_NOTEX] [-rt REPLICATES_TEX]
[-rf REPLICATES_FRAG] [-tb] [-ml MIN_LOOP_LENGTH]
[-Ml MAX_LOOP_LENGTH] [-ms MIN_STEM_LENGTH]
[-Ms MAX_STEM_LENGTH] [-mr MISS_RATE]
[-mu MIN_U_TAIL_LENGTH] [-ru RANGE_U_TAIL]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--TransTermHP_path TRANSTERMHP_PATH
Please assign the path of "transterm" in TransTermHP.
--expterm_path EXPTERM_PATH
Please assign the path of your expterm.dat for
TransTermHP. Default is /usr/local/bin/expterm.dat
--RNAfold_path RNAFOLD_PATH
If you want to assign the path of "RNAfold" of Vienna
package, please assign here.
--fasta_folder FASTA_FOLDER, -f FASTA_FOLDER
The path of genome fasta folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder.
--transcript_folder TRANSCRIPT_FOLDER, -a TRANSCRIPT_FOLDER
The folder which stores gff files of transcript
assembly.
--sRNA SRNA, -sr SRNA
If you want to include sRNA information, please assign
the folder of sRNA gff files.
--statistics, -s Doing statistics for TransTermHP. The name of
statistics file is - stat_terminator_$STRAIN_NAME.csv.
Default is False.
--tex_wig_folder TEX_WIG_FOLDER, -tw TEX_WIG_FOLDER
If you want to use tex +/- libraries, please assign
tex +/- wig folder.
--frag_wig_folder FRAG_WIG_FOLDER, -fw FRAG_WIG_FOLDER
If you want to use fragmented libraries, please assign
fragmented wig folder.
--decrease DECREASE, -d DECREASE
If the (lowest coverage / highest coverage) in the
terminator is smaller than this number, it will
consider this terminator have coverage dramatic
decreasing in it. Default is 0.5.
--fuzzy_detect_coverage FUZZY_DETECT_COVERAGE, -fc FUZZY_DETECT_COVERAGE
It will elongate the number of nucleotides(you assign
here) from both terminal site. If it can found the
coverage dramatic decreasing within this range, it
will still consider the terminator have coverage
dramatic decrease in it. Default is 30.
--fuzzy_within_transcript FUZZY_WITHIN_TRANSCRIPT, -fut FUZZY_WITHIN_TRANSCRIPT
If the candidates are within transcript and the
distance between the end of gene/transcript and
terminator candidate is within this number, it will be
consider as terminator. Default is 30.
--fuzzy_downstream_transcript FUZZY_DOWNSTREAM_TRANSCRIPT, -fdt FUZZY_DOWNSTREAM_TRANSCRIPT
If the candidates are downstream of transcript and the
distance between the end of gene/transcript and
terminator candidate is within this number, it will be
consider as terminator. Default is 30.
--fuzzy_within_gene FUZZY_WITHIN_GENE, -fuc FUZZY_WITHIN_GENE
If the candidates are upstream of gene and the
distance between the end of gene and terminator
candidate is within this number, it will be consider
as terminator. Default is 10.
--fuzzy_downstream_gene FUZZY_DOWNSTREAM_GENE, -fdg FUZZY_DOWNSTREAM_GENE
If the candidates are downstream of gene and the
distance between the end of gene and terminator
candidate is within this number, it will be consider
as terminator. Default is 310.
--highest_coverage HIGHEST_COVERAGE, -hc HIGHEST_COVERAGE
If the highest coverage of terminator is below to this
number, the terminator will be classify to non-detect,
but still included in "all_candidates". Default is 10.
-tl TEX_NOTEX_LIBS, --tex_notex_libs TEX_NOTEX_LIBS
Library name of tex and notex library. The format is:
wig_file_name:tex_treat_or_not(tex or notex):condition
_id(integer):replicate_id(alphabet):strand(+ or -). If
you have multiple wig files, please use comma to
separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
-fl FRAG_LIBS, --frag_libs FRAG_LIBS
Library name of fragmented library. The format is: wig
_file_name:fragmented(frag):condition_id(integer):repl
icate_id(alphabet):strand(+ or -). If you have
multiple wig files, please use comma to separate the
wig files. For example,
wig1:frag:1:a:+,wig2:frag:1:a:-.
-te TEX_NOTEX, --tex_notex TEX_NOTEX
For tex +/- library, terminators should be detected by
both or just one.(1/2) Default is 1.
-rt REPLICATES_TEX, --replicates_tex REPLICATES_TEX
The terminator of tex +/- library should be detected by
more than this number of replicates.
-rf REPLICATES_FRAG, --replicates_frag REPLICATES_FRAG
The terminator of fragmented library should be
detected more by than this number of replicates.
-tb, --table_best Output table only contains most decreasing track.
Default is False.
-ml MIN_LOOP_LENGTH, --min_loop_length MIN_LOOP_LENGTH
The minimum length of loop for terminator. Default is
3 nts.
-Ml MAX_LOOP_LENGTH, --max_loop_length MAX_LOOP_LENGTH
The maximum length of loop for terminator. Default is
10 nts.
-ms MIN_STEM_LENGTH, --min_stem_length MIN_STEM_LENGTH
The minimum length of stem for terminator. Default is
4 nts.
-Ms MAX_STEM_LENGTH, --max_stem_length MAX_STEM_LENGTH
The maximum length of stem for terminator. Default is
20 nts.
-mr MISS_RATE, --miss_rate MISS_RATE
How many percentage of nucleotides which has no base
pair in the stem. Default is 0.25.
-mu MIN_U_TAIL_LENGTH, --min_U_tail_length MIN_U_TAIL_LENGTH
The minimum length of U tail for terminator. Default
is 3 nts.
-ru RANGE_U_TAIL, --range_U_tail RANGE_U_TAIL
How long of nucleotides that you want to detect for U
tail. For example, if --range_U_tail is 6 and
--min_U_tail_length is 3, and there are 3 U within 6
nts, it will be assigned to detecting U tail
successfully. Default is 6.
- Output files
The output files will stored in $ANNOgesic/output/terminator
.
statistics
: Statistics files.
transtermhp
: All output of TranstermHP.
gffs
: Gff files of terminator.
There are four different sub-folders to store terminators.
all_candidates
will store all terminators which ANNOgesic
can detect.
express
will store the terminators which has gene expression.
best
will store the terminators which not only has gene expression but also
has coverage dramatic decreasing.
non_express
will store the terminators which has no gene expression.
tables
: The tables of terminators which store more details.
The tags of gff files:
Method
: The method that this terminator be detected.
coverage_decrease
: The coverage of the terminator has dramatic decreasing or not.
express
: The terminator has gene expression or not.
diff_coverage
: The highest coverage and lowest coverage of the library which expresses highest.
The numbers in parens are highest coverage and lowest coverage.
utr¶
utr
can compare with the information of TSSs, CDSs/tRNAs/sRNAs, transcripts and terminators
to generate proper UTRs. 5’UTRs are based on detecting the regions between TSSs (which
are located in transcripts) and CDSs/tRNAs/sRNAs. 3’UTRs are based on detecting the
regions between the end of transcripts and CDSs/tRNAs/sRNAs. If the gff files of TSSs are not computed by
ANNOgesic, please use –TSS_source. utr
would classify TSSs for the analysis.
- Pre-required files
Gff files of genome annotations, TSSs and transcripts.
If user wants to combine the information of terminators, it also need the gff files of terminator.
- Arguments
usage: annogesic utr [-h] [--annotation_folder ANNOTATION_FOLDER]
[--TSS_folder TSS_FOLDER]
[--transcript_assembly_folder TRANSCRIPT_ASSEMBLY_FOLDER]
[--terminator_folder TERMINATOR_FOLDER] [--TSS_source]
[--base_5UTR BASE_5UTR] [--UTR_length UTR_LENGTH]
[--base_3UTR BASE_3UTR]
[--terminator_fuzzy TERMINATOR_FUZZY]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder.
--TSS_folder TSS_FOLDER, -t TSS_FOLDER
The path of TSS folder.
--transcript_assembly_folder TRANSCRIPT_ASSEMBLY_FOLDER, -a TRANSCRIPT_ASSEMBLY_FOLDER
The path of transcriptome assembly folder.
--terminator_folder TERMINATOR_FOLDER, -e TERMINATOR_FOLDER
If you want to add the information of terminator, you
can assign the path of terminator folder here.
--TSS_source, -s If you generate TSS which is not from ANNOgesic,
please turn it on. Default is True(ANNOgesic).
--base_5UTR BASE_5UTR, -b5 BASE_5UTR
Which information that you want to use for generating
5'UTR. TSS/transcript/both. Default is both.
--UTR_length UTR_LENGTH, -l UTR_LENGTH
The maximum length of UTR. Default is 300.
--base_3UTR BASE_3UTR, -b3 BASE_3UTR
Which information that you want to use for generating
3'UTR. transcript/terminantor/both. Default is
transcript.
--terminator_fuzzy TERMINATOR_FUZZY, -f TERMINATOR_FUZZY
This is only for --base_3UTR which assigned by
"transcript" or "both". If the distance(nucleotides)
between terminator and the end of transcript lower
than this value, it will assign the terminator
associated with the 3'UTR. Default is 30.
- Output files
All output of 5’UTRs will store in $ANNOgesic/output/UTR/5UTR
.
All output of 3’UTRs will store in $ANNOgesic/output/UTR/3UTR
.
gffs
: Gff files of 5’UTR/3’UTR
The tags of gff files:
length
: UTR length.
associated_cds
: Which CDSs/rRNAs/tRNAs are associated with this UTR.
associated_gene
: Which genes are associated with this UTR.
associated_tran
: Which transcript is associated with this UTR.
associated_tss
: Which TSSs are associated with this 5’UTR.
TSS_type
: What types of TSSs are associated with this 5’UTR.
associated_term
: Which terminators are associated with this 3’UTR.
srna¶
srna
can predict sRNAs through comparing the transcripts and annotation profile.
The transcripts in intergenic region might be sRNA candidates. Moreover, based on
the information of TSSs and processing sites, we can also predict UTR-derived sRNA candidates.
- Pre-required tools and files
sRNA database, such as BSRD.
Gff files of genome annotation and Transcript data.
It can also import more information to improve the accuracy of prediction.
wiggle files: The libraries and wiggle files, Please refer to the The format of libraries for import to ANNOgesic
.
If user want to detect the UTR-derived sRNAs, it will be necessary to import more information.
TSSs: UTR-derived sRNAs must to generated from TSSs. If you don’t want to detect UTR-derived sRNAs, you can also import this information for statistics.
processing sites: It is for check the sRNAs which end with processing sites. Moreover, Some 3’UTR-derived and interCDS-derived sRNA candidates start from processing sites not TSSs. If you don’t want to detect UTR-derived sRNAs, you can also import this information for statistics.
Optional input file:
Fasta files of genome sequence.
sRNA database: It could be used for searching the known sRNA. The format of header should be
$ID|$STRAIN|$SRNANAME
.
For example, >saci2813.1|Acinetobacter sp. ADP1|Aar
.
The ID is saci403.1; the strain of this sRNA is Acinetobacter sp. ADP1 and the name of sRNA is Aar.
If the user doesn’t follow the format, it will occur error when the user runs with --sRNA_blast_stat, -sb
.
Or the results will be meaningless.
Of course, it can run srna
without --sRNA_blast_stat, -sb
.
nr database: It could be used for searching the known proteins in order to exclude false positive.
Gff files of sORFs: It could compare sORFs and sRNAs. User can refer these information to find the best sRNA candidates.
Gff files of terminator: It could compare terminators and sRNAs. User can refer these information to find the best sRNA candidates.
Table of promoters: It could compare promoters and sRNAs. User can refer these information to find the best sRNA candidates. The format should be
| strain | TSS_position | TSS_strand | Motif |
| NC_000915.1 | 237118 | - | MOTIF_1 |
| NC_000915.1 | 729009 | - | MOTIF_1 |
First row is the header of table, the last column is the name of motif/promoter.
If you want apply ANNOgesci to compute the promoters, the table will be generated automatically.
Please refer to the section of promoter
.
 If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--Vienna_folder VIENNA_FOLDER
Please assign the folder of Vienna package. It should
include RNAfold.
--Vienna_utils VIENNA_UTILS
Please assign the folder of Utils of Vienna package.
It should include relplot.pl and mountain.pl.
--blast_plus_folder BLAST_PLUS_FOLDER
Please assign the folder of blast+ which include
blastn, blastx, makeblastdb.
--ps2pdf14_path PS2PDF14_PATH
Please assign the path of ps2pdf14.
--UTR_derived_sRNA, -u
If you want to detect UTR-derived sRNA, please turn it
on. Default is False.
--import_info IMPORT_INFO, -d IMPORT_INFO
There are several types of information that you can
import to detect and filter sRNAs: tss(the sRNA should
start from a TSS), sec_str(free energy change of
secondary structure(normalized by length)),
blast_nr(blast to nr), blast_srna(blast to sRNA),
sorf(compare with sORF), term(compare with
terminator), promoter(compare with promoter motif).
ATTENTION: without filters, the results may be not
well. Please assign the information you want to import
(comma for separate the filters), i.e.
tss,sec_str,blast_nr - means it used TSS, energy and
blast result to detect sRNA. Besides these
information, it will also consider the sequence length
of sRNA. ATTENTION: if you want to import sRNA
database, please follow the format that we define
$ID|$STRAIN|$SRNANAME. Default is
tss,sec_str,blast_nr,blast_srna.
--transcript_assembly_folder TRANSCRIPT_ASSEMBLY_FOLDER, -a TRANSCRIPT_ASSEMBLY_FOLDER
The path of transcriptome assembly folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder.
--TSS_folder TSS_FOLDER, -t TSS_FOLDER
If you want to import TSS information, please assign
the path of gff folder of TSS. If you want to detect
UTR-derived sRNA, you MUST assign the folder of TSS.
--processing_site_folder PROCESSING_SITE_FOLDER, -p PROCESSING_SITE_FOLDER
If you want to import processing site information,
please assign the path of gff folder of processing
site.If you want to detect UTR-derived sRNA, you MUST
assign the folder of processing site.
--promoter_table PROMOTER_TABLE, -pt PROMOTER_TABLE
If you want to import promoter information, please
assign the path of promoter table. The format of table
is $STRAIN $TSS_POSITION $TSS_STRAND $PROMOTER_NAME.
If you want to import promoter information, the
associated tss imformation is required.
--promoter_name PROMOTER_NAME, -pn PROMOTER_NAME
If you want to import promoter information, please
assign the promoter name (the last column of promoter
table) which you want to compare. If you want to
import multiple promoters, please put comma between
the promoters. Default is None.
--TSS_source, -ts If your gff file of TSS is not generated by ANNOgesic,
please you turn it on. It will classify TSSs and the
proper format for sRNA prediction. Default is True.
--TSS_intergenic_fuzzy TSS_INTERGENIC_FUZZY, -ft TSS_INTERGENIC_FUZZY
If you want to import TSS information, you need to
assign the fuzzy for comparing TSS and transcript
assembly/CDS. It is for intergenic.Default is 3.
--TSS_5UTR_fuzzy TSS_5UTR_FUZZY, -f5 TSS_5UTR_FUZZY
If you want to import TSS information, you need to
assign the fuzzy for comparing TSS and transcript
assembly. It is for 5'UTR of UTR derived sRNA.You can
use percentage or the amount of reads. p_0.05 means
the fuzzy is 5 percent of the length of CDS which
overlap with transcript. n_10 means the fuzzy is 10
base pair. Default is n_3.
--TSS_3UTR_fuzzy TSS_3UTR_FUZZY, -f3 TSS_3UTR_FUZZY
If you want to import TSS information, you need to
assign the fuzzy for comparing TSS and transcript
assembly/CDS. It is for 3'UTR of UTR derived sRNA.You
can use percentage or the amount of reads. p_0.05
means the fuzzy is 5 percent of the length of CDS
which overlap with transcript. n_10 means the fuzzy is
10 base pair. Default is p_0.04.
--TSS_interCDS_fuzzy TSS_INTERCDS_FUZZY, -fc TSS_INTERCDS_FUZZY
If you want to import TSS information, you need to
assign the fuzzy for comparing TSS and transcript
assembly. It is for interCDS derived sRNA.You can use
percentage or the amount of reads. p_0.05 means the
fuzzy is 5 percent of the length of CDS which overlap
with transcript. n_10 means the fuzzy is 10 base pair.
Default is p_0.04.
--terminator_folder TERMINATOR_FOLDER, -tf TERMINATOR_FOLDER
If you want to import terminator information, please
assign the path of gff folder of terminator.
--terminator_fuzzy_in_CDS TERMINATOR_FUZZY_IN_CDS, -tfi TERMINATOR_FUZZY_IN_CDS
If you want to import terminator information, you need
to assign the fuzzy for comparing terminator and
transcript assembly. It is the fuzzy for the
terminator which is within CDS. Default is 30.
--terminator_fuzzy_out_CDS TERMINATOR_FUZZY_OUT_CDS, -tfo TERMINATOR_FUZZY_OUT_CDS
If you want to import terminator information, you need
to assign the fuzzy for comparing terminator and
transcript assembly. It is the fuzzy for the
terminator which is out of CDS. Default is 30.
--min_length MIN_LENGTH, -lm MIN_LENGTH
Please assign the minimum length of sRNA. Default
is 30.
--max_length MAX_LENGTH, -lM MAX_LENGTH
Please assign the maximum length of sRNA. Default
is 500.
--tex_wig_folder TEX_WIG_FOLDER, -tw TEX_WIG_FOLDER
The path of tex+/- wig folder.
--frag_wig_folder FRAG_WIG_FOLDER, -fw FRAG_WIG_FOLDER
The path of fragment wig folder.
--run_intergenic_TEX_coverage RUN_INTERGENIC_TEX_COVERAGE, -it RUN_INTERGENIC_TEX_COVERAGE
The minimum average coverage of intergenic sRNA
candidates for TEX +. You can choose what kinds of TSS
that you want to use for sRNA prediction and assign
the cutoff of coverage to TSS. The order of numbers is
"Primary Secondary Internal Antisense Orphan"
(separated by comma). Ex: if you assign 0,0,0,50,10,
it means you want to use Antisense(cutoff coverage is
50) and Orphan(cutoff coverage is 10) for sRNA
prediction. 0 means you don't want to use it for
prediction. If you don't use TSS information for
prediction, it will choose the lowest one as a cutoff.
Ex: if you assign 0,0,0,50,10 and you have no TSS
information, it will use 10 as a general cutoff for
prediction. Default is 0,0,0,40,20.
--run_intergenic_noTEX_coverage RUN_INTERGENIC_NOTEX_COVERAGE, -in RUN_INTERGENIC_NOTEX_COVERAGE
The minimum average coverage of intergenic sRNA
candidates for TEX -. You can choose what kinds of TSS
that you want to use for sRNA prediction and assign
the cutoff of coverage to TSS. The order of numbers is
"Primary Secondary Internal Antisense Orphan"
(separated by comma). Ex: if you assign 0,0,0,50,10,
it means you want to use Antisense(cutoff coverage is
50) and Orphan(cutoff coverage is 10) for sRNA
prediction. 0 means you don't want to use it for
prediction. If you don't use TSS information for
prediction, it will choose the lowest one as a cutoff.
Ex: if you assign 0,0,0,50,10 and you have no TSS
information, it will use 10 as a general cutoff for
prediction. Default is 0,0,0,30,10.
--run_intergenic_fragmented_coverage RUN_INTERGENIC_FRAGMENTED_COVERAGE, -if RUN_INTERGENIC_FRAGMENTED_COVERAGE
The minimum average coverage of intergenic sRNA
candidates for fragmented library. You can choose what
kinds of TSS that you want to use for sRNA prediction
and assign the cutoff of coverage to TSS. The order of
numbers is "Primary Secondary Internal Antisense
Orphan" (separated by comma). Ex: if you assign
0,0,0,50,10, it means you want to use Antisense(cutoff
coverage is 50) and Orphan(cutoff coverage is 10) for
sRNA prediction. 0 means you don't want to use it for
prediction. If you don't use TSS information for
prediction, it will choose the lowest one as a cutoff.
Ex: if you assign 0,0,0,50,10 and you have no TSS
information, it will use 10 as a general cutoff for
prediction. Default is 400,200,0,50,20.
--run_antisense_TEX_coverage RUN_ANTISENSE_TEX_COVERAGE, -at RUN_ANTISENSE_TEX_COVERAGE
The meaning is the same as
--run_intergenic_TEX_coverage. Just apply to
antisense. Default is 0,0,0,40,20.
--run_antisense_noTEX_coverage RUN_ANTISENSE_NOTEX_COVERAGE, -an RUN_ANTISENSE_NOTEX_COVERAGE
The meaning is the same as
--run_intergenic_noTEX_coverage. Just apply to
antisense. Default is 0,0,0,30,10.
--run_antisense_fragmented_coverage RUN_ANTISENSE_FRAGMENTED_COVERAGE, -af RUN_ANTISENSE_FRAGMENTED_COVERAGE
The meaning is the same as
--run_intergenic_fragmented_coverage. Just apply to
antisense. Default is 400,200,0,50,20.
--intergenic_tolerance INTERGENIC_TOLERANCE, -ti INTERGENIC_TOLERANCE
This number indicates the tolerance of temporary drop
below cutoff of coverage. Default is 5.
--run_utr_TEX_coverage RUN_UTR_TEX_COVERAGE, -ut RUN_UTR_TEX_COVERAGE
The minimum average coverage of UTR-derived sRNA
candidates for TEX +. You can assign the percentile or
real number of coverage for 5'UTR, 3'UTR and interCDS.
The order of numbers are "5'UTR, 3'UTR and interCDS"
(separated by comma). Ex: if you assign
"p_0.7,p_0.5,p_0.5", it will use 70 percentile of
coverage as cutoff for 5'UTR, median of coverage as
cutoff for 3'UTR and interCDS. Ex: if you assign
"n_30,n_10,n_20 " it will use 30 as cutoff for 5'UTR
and 10 as cutoff for 3'UTR and 20 for interCDS.
Default is p_0.8,p_0.6,p_0.7.
--run_utr_noTEX_coverage RUN_UTR_NOTEX_COVERAGE, -un RUN_UTR_NOTEX_COVERAGE
The minimum average coverage of UTR-derived sRNA
candidates for TEX -. You can assign the percentile or
real number of coverage for 5'UTR, 3'UTR and interCDS.
The order of numbers are "5'UTR, 3'UTR and interCDS"
(separated by comma). Ex: if you assign
"p_0.7,p_0.5,p_0.5", it will use 70 percentile of
coverage as cutoff for 5'UTR, median of coverage as
cutoff for 3'UTR and interCDS. Ex: if you assign
"n_30,n_10,n_20 " it will use 30 as cutoff for 5'UTR
and 10 as cutoff for 3'UTR and 20 for interCDS.
Default is p_0.7,p_0.5,p_0.6.
--run_utr_fragmented_coverage RUN_UTR_FRAGMENTED_COVERAGE, -uf RUN_UTR_FRAGMENTED_COVERAGE
The minimum average coverage of UTR-derived sRNA
candidates for fragmented library. You can assign the
percentile or real number of coverage for 5'UTR, 3'UTR
and interCDS. The order of numbers are "5'UTR, 3'UTR
and interCDS" (separated by comma). Ex: if you assign
"p_0.7,p_0.5,p_0.5", it will use 70 percentile of
coverage as cutoff for 5'UTR, median of coverage as
cutoff for 3'UTR and interCDS. Ex: if you assign
"n_30,n_10,n_20 " it will use 30 as cutoff for 5'UTR
and 10 as cutoff for 3'UTR and 20 for interCDS.
Default is p_0.7,p_0.5,p_0.6.
--min_utr_coverage MIN_UTR_COVERAGE, -mu MIN_UTR_COVERAGE
The minimum coverage of UTR-derived sRNA. The coverage
should not only fit the --run_utr_TEX_coverage,
--run_utr_noTEX_coverage and
--run_utr_fragmented_coverage, but also this value.
Defaul is 50.
--fasta_folder FASTA_FOLDER, -f FASTA_FOLDER
If you want to import sec_str, blast_nr, blast_srna
(--import_info), please assign the path of genome
fasta folder.
--cutoff_energy CUTOFF_ENERGY, -e CUTOFF_ENERGY
If you want to import secondary structure information,
please assign the cutoff of folding energy change
(normalized by length of gene). Default is -0.05.
--mountain_plot, -m If you want to generate mountain plots of sRNA
candidates, please turn it on. Default is False.
--nr_format, -nf It is for formating nr database. If you already format
nr database, you don't need to turn it on. Default is
False.
--srna_format, -sf It is for formating sRNA database. If you already
format sRNA database, you don't need to turn it on.
Default is False.
--sRNA_database_path SRNA_DATABASE_PATH, -sd SRNA_DATABASE_PATH
If you want to import blast results of sRNA, please
assign the path of sRNA database.
--nr_database_path NR_DATABASE_PATH, -nd NR_DATABASE_PATH
If you want to import blast results of nr, please
assign the path of nr database.
--tex_notex_libs TEX_NOTEX_LIBS, -tl TEX_NOTEX_LIBS
library name of tex and notex libraries. The format
is: wig_file_name:tex_treat_or_not(tex or notex):condi
tion_id(integer):replicate_id(alphabet):strand(+ or
-). If you have multiple wig files, please use comma
to separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--frag_libs FRAG_LIBS, -fl FRAG_LIBS
library name of fragmented libraries. The format is: w
ig_file_name:fragmented(frag):condition_id(integer):re
plicate_id(alphabet):strand(+ or -). If you have
multiple wig files, please use comma to separate the
wig files. For example,
wig1:frag:1:a:+,wig2:frag:1:a:-.
--tex_notex TEX_NOTEX, -te TEX_NOTEX
For tex +/- libraries, sRNA candidates should be
detected by both or just one.(1/2) Default is 2.
--replicates_tex REPLICATES_TEX, -rt REPLICATES_TEX
The sRNA of tex +/- libraries should be detected by
more than this number of replicates.
--replicates_frag REPLICATES_FRAG, -rf REPLICATES_FRAG
The sRNA of fragmented libraries should be detected by
more than this number of replicates.
--table_best, -tb The output table of sRNA candidates only print the
best track of libraries. Default is False.
--decrease_intergenic DECREASE_INTERGENIC, -di DECREASE_INTERGENIC
If the intergenic region is longer than the
max_length, it will based on coverage to check the
sRNA candidates. If (lowest coverage / the highest
coverage) of intergenic region is smaller than this
number, it will consider the the spot of lowest
coverage as end point. If the length of from start to
the end point is proper for sRNA candidate, it also
consider it as a sRNA candidate. Default is 0.1.
--decrease_utr DECREASE_UTR, -du DECREASE_UTR
It is similar with --decrease_intergenic. It is for
UTR-derived sRNAs. Default is 0.05.
--fuzzy_intergenic FUZZY_INTERGENIC, -fi FUZZY_INTERGENIC
If the situation is like --decrease_intergenic
mentioned, it is fuzzy value between the end of sRNA.
Default is 10.
--fuzzy_utr FUZZY_UTR, -fu FUZZY_UTR
It is simliar with --fuzzy_intergenic. It is for UTR-
derived sRNAs. Default is 10.
--cutoff_nr_hit CUTOFF_NR_HIT, -cn CUTOFF_NR_HIT
The cutoff of number of hits in nr database. If the
number of nr hits more than this cutoff, it will be
excluded. Default is 0.
--blast_e_nr BLAST_E_NR, -en BLAST_E_NR
The cutoff of blast e value for nr alignment. Default
is 0.0001.
--blast_e_srna BLAST_E_SRNA, -es BLAST_E_SRNA
The cutoff of blast e value for sRNA alignment.
Default is 0.0001.
--sORF SORF, -O SORF If you want to compare sORF and sRNA, please assign
the path of sORF gff folder.
--best_with_all_sRNAhit, -ba
When you want to include the sRNA candidates which can
find the homology from blast sRNA database without
considering other information(ex. TSS, blast in nr...)
to the best results. Please turn it on. Or it will
just select the best candidates based on all filter
conditions. Default is False.
--best_without_sORF_candidate, -bs
If you want to generate the best sRNA candidates which
excluded the overlap with sORFs.Please turn it on. Or
it will select the best candidates without considering
the overlap with sORF. Default is False.
--best_with_terminator, -bt
If you want to generate the best sRNA candidates which
must be associated with terminator. Please turn it on.
Or it will select the best candidates without
considering the terminator. If the sRNA candidate ends
with processing site, it will include to best results,
too. Default is False.
--best_with_promoter, -bp
If you want to generate the best sRNA candidates which
is must be associated with promoter.Please turn it on.
Default is False.
--detect_sRNA_in_CDS, -ds
If you assume there are some wrong annotation in your
genome annotation file and you want to search the
small transcript in CDS, you can turn it on. It may
find more sRNA candidates which overlap with CDS.
Default is False.
--overlap_percent_CDS OVERLAP_PERCENT_CDS, -oc OVERLAP_PERCENT_CDS
If you want to execute --detect_sRNA_in_CDS, you can
assign the cutoff. If the ratio of overlap between CDS
and sRNA candidates is lower than the cutoff, It may
be sRNA candidates. Default is 0.5
--ignore_hypothetical_protein, -ih
If you want to ignore hypothetical protein in genome
annotation file, you can turn it on. Default is False.
--ranking_time_promoter RANKING_TIME_PROMOTER, -rp RANKING_TIME_PROMOTER
If you imported promoter information, you can also use
it to rank sRNA candidates. The ranking will base on
--ranking_time_promoter * average coverage. For
example, one candidates which average coverage is 10,
associated with promoter and --ranking_time_promoter
is 2, the score for ranking will be 20 (2*10). The
candidates which are not associated with promoters,
the --ranking_time_promoter is 1. Default is 2. This
number can not be smaller than 1.
- Output files
All output files will be stored in $ANNOgesic/output/sRNA
.
sRNA_2d_$STRAIN_NAME
: The secondary structure of all sRNA candidates.
sRNA_seq_$STRAIN_NAME
: The sequence of all sRNA candidates.
blast_result_and_misc
: The results of blast.
mountain_plot
: The mountain plots of sRNA candidates.
sec_structure
: The dot plots and secondary structure plots of sRNA candidates.
statistics
: Statistics files. stat_$STRAIN_NAME_sRNA_blast.csv
is the results of analysis of blast sRNA databases.
stat_sRNA_class_Staphylococcus_aureus_HG003.csv
is the results of classification of sRNA candidates.
tables
: sRNA tables for more details. It also includes the ranking of sRNA candidates.
for class
is for different classes of sRNAs.
best
is the best results of sRNAs after filtering. all_candidates
is for all candidates without filtering.
gffs
: Gff files of sRNAs. The meanings of for class
, best
, all_candidates
are the same as tables
.
The useful tags of gff files:
sRNA_type
: The sRNA is from 5’UTR or 3’UTR or interCDS or intergenic or antisense or in_CDS.
with_TSS
: Which TSSs are related to this sRNA. NA means the sRNA is not related to any TSSs.
sORF
: Which sORFs overlap with this sRNA.
sRNA_hit
: The blast hit of sRNA database.
nr_hit
: The blast hit of nr database.
2d_energy
: The normalized(by the length of sRNA) free energy change of secondary structure of sRNA candidate.
with_term
: The terminators which associated with the sRNA candidate.
If you assigned --TSS_source
for sRNA prediction, TSS_class
will be generated and store the gff files of TSSs.
promoter
: The promoters which associated with this sRNA candidates.
overlap_cds
: The CDSs which overlap with sRNA.
overlap_percent
: If there are CDSs overlap with sRNA, it will shows the percentage of overlap.
sorf¶
sorf
can detect sORF based on ribosomal binding sites, start codons and stop codons within the intergenic transcripts.
Because non-coding region may be sRNAs or sORFs, it also provide the function to compare sORFs and sRNAs.
If there are some sORFs overlaped, it will merge them together. Therefore, one region may contain more than one sORF.
BE CAREFUL, The position of start codon is the first nucleotide. The position of stop codon is the last nucleotide.
As we know that three nucleotides form one amino acid. Therefore, one small transcript may contain
three open reading frames for one strand. Since sorf
provides the region which covers all possible sORFs,
the start point and end point may not be the multiple of 3. The region may contain different frame shift.
Ex: (200, 202, 203) are the positions of three start codons and (241, 243) are two stop codons in
a small transcript. We can see that there are three possible ORFs(200-241, 203-241 and 202-243).
for 200-241 and 203-241, the frame has no shift, because the difference of positions for these two
ORFs are the multiple of 3. However, for 202-243, there is a frame shift. Please be aware this point for
using the results.
- Pre-required tools and files
The gff files of CDSs/tRNAs/rRNAs and transcripts.
The libraries and wiggle files, Please refer to the The format of libraries for import to ANNOgesic
.
The fasta files of genome sequence for detection of ribosomal binding sites, start codons and stop codons.
User can also import some useful information to improve the prediction:
gff files of TSSs for checking the sORFs start from TSS or not.
gff files of sRNAs for checking the conflict of sRNAs and sORFs.
- Arguments
usage: annogesic sorf [-h] [--UTR_derived_sORF]
[--transcript_assembly_folder TRANSCRIPT_ASSEMBLY_FOLDER]
[--annotation_folder ANNOTATION_FOLDER]
[--TSS_folder TSS_FOLDER] [--utr_length UTR_LENGTH]
[--min_length MIN_LENGTH] [--max_length MAX_LENGTH]
[--tex_wig_folder TEX_WIG_FOLDER]
[--frag_wig_folder FRAG_WIG_FOLDER]
[--cutoff_intergenic_coverage CUTOFF_INTERGENIC_COVERAGE]
[--cutoff_antisense_coverage CUTOFF_ANTISENSE_COVERAGE]
[--cutoff_5utr_coverage CUTOFF_5UTR_COVERAGE]
[--cutoff_3utr_coverage CUTOFF_3UTR_COVERAGE]
[--cutoff_interCDS_coverage CUTOFF_INTERCDS_COVERAGE]
[--cutoff_background CUTOFF_BACKGROUND]
[--fasta_folder FASTA_FOLDER]
[--tex_notex_libs TEX_NOTEX_LIBS]
[--frag_libs FRAG_LIBS] [--tex_notex TEX_NOTEX]
[--replicates_tex REPLICATES_TEX]
[--replicates_frag REPLICATES_FRAG] [--table_best]
[--sRNA_folder SRNA_FOLDER] [--start_codon START_CODON]
[--stop_codon STOP_CODON]
[--min_rbs_distance MIN_RBS_DISTANCE]
[--max_rbs_distance MAX_RBS_DISTANCE]
[--rbs_not_after_TSS] [--fuzzy_rbs FUZZY_RBS]
[--print_all_combination] [--best_no_sRNA]
[--best_no_TSS]
[--ignore_hypothetical_protein IGNORE_HYPOTHETICAL_PROTEIN]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--UTR_derived_sORF, -u
If you want to detect UTR-derived sORF, please turn it
on. Default is False.
--transcript_assembly_folder TRANSCRIPT_ASSEMBLY_FOLDER, -a TRANSCRIPT_ASSEMBLY_FOLDER
The path of transcriptome assembly folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder.
--TSS_folder TSS_FOLDER, -t TSS_FOLDER
If you want to import TSS information, please assign
the path of gff folder of TSS.
--utr_length UTR_LENGTH, -ul UTR_LENGTH
If you want to import TSS information, please assign
the utr length for comparing TSS and sORF. The default
number is 300.
--min_length MIN_LENGTH, -lm MIN_LENGTH
Please assign the minimum length of sORF. It will
predict sORF candidates based on the value. Default is
30.
--max_length MAX_LENGTH, -lM MAX_LENGTH
Please assign the maximum length of sORF. It will
predict sORF candidates based on the value. Default is
150.
--tex_wig_folder TEX_WIG_FOLDER, -tw TEX_WIG_FOLDER
The path of tex+/- wig folder.
--frag_wig_folder FRAG_WIG_FOLDER, -fw FRAG_WIG_FOLDER
The path of fragment wig folder.
--cutoff_intergenic_coverage CUTOFF_INTERGENIC_COVERAGE, -ci CUTOFF_INTERGENIC_COVERAGE
The cutoff of minimum coverage of intergenic sORF
candidates.
--cutoff_antisense_coverage CUTOFF_ANTISENSE_COVERAGE, -ai CUTOFF_ANTISENSE_COVERAGE
The cutoff of minimum coverage of antisense sORF
candidates.
--cutoff_5utr_coverage CUTOFF_5UTR_COVERAGE, -cu5 CUTOFF_5UTR_COVERAGE
The cutoff of minimum coverage of 5'UTR derived sORF
candidates. You can use percentage or the amount of
reads. p_0.05 means the coverage of sORF candidates
should higher than 5 percentile of all 5'UTR
transcripts. n_10 means the coverage of sORF
candidates should be 10. Default is p_0.5.
--cutoff_3utr_coverage CUTOFF_3UTR_COVERAGE, -cu3 CUTOFF_3UTR_COVERAGE
The cutoff of minimum coverage of 3'UTR derived sORF
candidates. You can use percentage or the amount of
reads. p_0.05 means the coverage of sORF candidates
should higher than 5 percentile of all 3'UTR
transcripts. n_10 means the coverage of sORF
candidates should be 10. Default is p_0.5.
--cutoff_interCDS_coverage CUTOFF_INTERCDS_COVERAGE, -cuf CUTOFF_INTERCDS_COVERAGE
The cutoff of minimum coverage of interCDS derived
sORF candidates. You can use percentage or the amount
of reads. p_0.05 means the coverage of sORF candidates
should higher than 5 percentile of all interCDS
transcripts. n_10 means the coverage of sORF
candidates should be 10. Default is p_0.5.
--cutoff_background CUTOFF_BACKGROUND, -cub CUTOFF_BACKGROUND
The cutoff of minimum coverage of all sORF candidates.
Default is 10.
--fasta_folder FASTA_FOLDER, -f FASTA_FOLDER
The folder of genome fasta file.
--tex_notex_libs TEX_NOTEX_LIBS, -tl TEX_NOTEX_LIBS
Library name of tex+/- library. The format is:
wig_file_name:tex_treat_or_not(tex or notex):condition
_id(integer):replicate_id(alphabet):strand(+ or -). If
you have multiple wig files, please use comma to
separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--frag_libs FRAG_LIBS, -fl FRAG_LIBS
Library name of fragmented library The format is: wig_
file_name:fragmented(frag):condition_id(integer):repli
cate_id(alphabet):strand(+ or -). If you have multiple
wig files, please use comma to separate the wig files.
For example, wig1:frag:1:a:+,wig2:frag:1:a:-.
--tex_notex TEX_NOTEX, -te TEX_NOTEX
For tex +/- library, sORF candidates should be
detected by both or just one.(1/2) Default is 2.
--replicates_tex REPLICATES_TEX, -rt REPLICATES_TEX
The sORF of tex +/- library should be detected by more
than this number of replicates.
--replicates_frag REPLICATES_FRAG, -rf REPLICATES_FRAG
The sORF of fragmented library should be detected by
more than this number of replicates.
--table_best, -tb The output table of sORF candidates only print the
best track. Default is False.
--sRNA_folder SRNA_FOLDER, -s SRNA_FOLDER
If you want to compare sORF and sRNA, please assign
the path of sORF gff folder.
--start_codon START_CODON, -ac START_CODON
What kinds of start coden you want to use. If you want
to assign more than one type of start codon, please
use comma to separate them. Default is ATG.
--stop_codon STOP_CODON, -oc STOP_CODON
What kinds of stop coden you want to use. If you want
to assign more than one type of stop codon, please use
comma to separate them. Default is TTA,TAG,TGA.
--min_rbs_distance MIN_RBS_DISTANCE, -mr MIN_RBS_DISTANCE
The minimum distance between the ribosomal binding
site and start codon. Default is 3.
--max_rbs_distance MAX_RBS_DISTANCE, -Mr MAX_RBS_DISTANCE
The maximum distance between the ribosomal binding
site and start codon. Default is 15.
--rbs_not_after_TSS, -at
If you want to generate best gff file which include
ribosomal binding site not after TSS, please turn it
on. Default is False.
--fuzzy_rbs FUZZY_RBS, -zr FUZZY_RBS
How many nucleotides of ribosomal binding site is
different with AGGAGG? Default is 2.
--print_all_combination, -pa
Every expressed transcript of sORF has many start
codons and stop codons. If you want to print all
combinations of start codons and stop codons, please
turn it on. Default is False.
--best_no_sRNA, -bs If you want to generate best gff file without
overlaping with sRNA, please turn it on. Default is
False.
--best_no_TSS, -bt If you want to generate best gff file which not refere
to TSS, please turn it on. Default is False.
--ignore_hypothetical_protein IGNORE_HYPOTHETICAL_PROTEIN, -ih IGNORE_HYPOTHETICAL_PROTEIN
If you want to ignore hypothetical protein in genome
annotation file, you can turn it on. Default is False.
- Output files
All output files will be stored in $ANNOgesic/output/sORF
.
statistics
: Statistics files.
tables
: The tables of sORFs for more details. all_candidates
is for all sORF candidates without filtering.
best
is for the best sORF candidates with filtering.
gffs
: Gff files of sORFs. The meanings of all_candidates
and best
are the same as tables
.
The tags of gff files:
start_TSS
: The starting TSS of this sORF.
with_TSS
: Which TSSs are associated with this sORFs.
sORF_type
: The type of the sORF (5’UTR, 3’UTR, interCDS, intergenic, antisense or in_CDS).
sRNA
: Which sRNAs overlap with the the region of sORFs.
RBS
: The ribosomal binding sites of the region of sORFs.
frame_shift
: How many frame shifts in the regions of sORFs.
promoter¶
promoter
can scan the upstream of TSSs to discover the promoter motifs.
User can assign the regions of upstream TSSs. We integrate MEME to compute the promoters.
User can view the results very easily. If the gff files of TSSs is not computed by
ANNOgesic, please use –TSS_source. promoter
will classify the TSSs for computing
promoter motifs.
- Pre-required tools and files
MEME.
Fasta files of genome sequence.
Gff files of genome annotation.
Gff files of TSSs.
If the gff files of TSSs is not computed by ANNOgesic, the libraries and wiggle files are necessary.
Please refer to the The format of libraries for import to ANNOgesic
in order to assign the correct format.
- Arguments
usage: annogesic promoter [-h] [--MEME_path MEME_PATH]
[--fasta_folder FASTA_FOLDER]
[--TSS_folder TSS_FOLDER] [--num_motif NUM_MOTIF]
[--nt_before_TSS NT_BEFORE_TSS] [--e_value E_VALUE]
[--motif_width MOTIF_WIDTH] [--TSS_source]
[--tex_libs TEX_LIBS] [--tex_wig_path TEX_WIG_PATH]
[--annotation_folder ANNOTATION_FOLDER]
[--combine_all]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--MEME_path MEME_PATH
path of MEME.
--fasta_folder FASTA_FOLDER, -f FASTA_FOLDER
Please assign the folder of genome fasta file.
--TSS_folder TSS_FOLDER, -t TSS_FOLDER
The folder of TSS gff file.
--num_motif NUM_MOTIF, -n NUM_MOTIF
How many of motifs you want to produce? Default is 10.
--nt_before_TSS NT_BEFORE_TSS, -b NT_BEFORE_TSS
How many nucleotides that you want to extract before
TSS for promoter prediction? Default is 50.
--e_value E_VALUE, -e E_VALUE
The cutoff of e value. Default is 0.05.
--motif_width MOTIF_WIDTH, -w MOTIF_WIDTH
Motif length - it will refer the value to find the
motif. If you want to detect a range of width, you can
insert "-" between two values. Moreover, If you want
to compute more than one motif lenght, please use
comma to separate them. for example, 50,2-10. It means
the range of width which you want to detect is 50 and
within 2 to 10. The number should be less or equal
than --nt_before_TSS. Default is 50.
--TSS_source, -s If you generate TSS from other method, please turn it
on. Default is True(from ANNOgesic)
--tex_libs TEX_LIBS, -tl TEX_LIBS
Library name of tex+/- library. If your TSS is not
from ANNOgesic, please assign it.The format is:
wig_file_name:tex_treat_or_not(tex or notex):condition
_id(integer):replicate_id(alphabet):strand(+ or -). If
you have multiple wig files, please use comma to
separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--tex_wig_path TEX_WIG_PATH, -tw TEX_WIG_PATH
The path of tex+/- wig folder. If your TSS is not from
ANNOgesic, please assign the folder of tex+/- wig
folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder. If your TSS
is not from ANNOgesic, please assign the annotation
gff path.
--combine_all, -c If you want to combine all TSS files in "TSS_folder"
to generate global promoter motifs, please turn it on.
Default is False.
- Output files
All output files will be stored in $ANNOgesic/output/promoter_analysis
.
allfasta
: If you turn --combine_all
on, it will combine all TSSs in --TSS_folder
to generate promoter motifs. The results will store in this folder.
fasta_class
: The fasta files of the classes of TSSs.
Every strain will generate one folder for storing the information of promoter motifs.
The sub-folder is promoter_motifs_$FILENAME_$STRAINNAME_$TSSTYPE_$PROMOTERLEGNTH
.
Each sub-folder stores PNG files and HTML files of promoters. It also contains a table(CSV file) of
prometer which can be used to sRNA detection (please refer to srna
).
If the TSSs are not computed by ANNOgesic, TSS_class
will be generated. It will classify the
TSSs and store as gff files in it.
operon¶
operon
will group TSSs, genes/CDSs/tRNAs/rRNAs, transcripts, terminators and UTRs to operons and
suboperons.
- Pre-required tools or files
Gff files of TSSs, annotations, transcripts, 5’UTRs, and 3’UTRs.
If user wants to import the information of terminators, operon
can integrate terminators, too.
- Arguments
usage: annogesic operon [-h] [--TSS_folder TSS_FOLDER]
[--annotation_folder ANNOTATION_FOLDER]
[--transcript_folder TRANSCRIPT_FOLDER]
[--UTR5_folder UTR5_FOLDER]
[--UTR3_folder UTR3_FOLDER]
[--term_folder TERM_FOLDER] [--TSS_fuzzy TSS_FUZZY]
[--term_fuzzy TERM_FUZZY] [--min_length MIN_LENGTH]
[--statistics] [--combine_gff]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--TSS_folder TSS_FOLDER, -t TSS_FOLDER
The path of TSS gff folder.
--annotation_folder ANNOTATION_FOLDER, -g ANNOTATION_FOLDER
The path of genome annotation gff folder.
--transcript_folder TRANSCRIPT_FOLDER, -a TRANSCRIPT_FOLDER
The path of transcript gff folder.
--UTR5_folder UTR5_FOLDER, -u5 UTR5_FOLDER
The path of 5'UTR gff folder.
--UTR3_folder UTR3_FOLDER, -u3 UTR3_FOLDER
The path of 3'UTR gff folder.
--term_folder TERM_FOLDER, -e TERM_FOLDER
If you want to import the information of terminator,
please assign the path of terminator gff folder.
--TSS_fuzzy TSS_FUZZY, -tf TSS_FUZZY
The fuzzy for comparing TSS and transcript assembly.
Default is 5.
--term_fuzzy TERM_FUZZY, -ef TERM_FUZZY
The fuzzy for comparing terminator and transcript
assembly. Default is 30.
--min_length MIN_LENGTH, -l MIN_LENGTH
The minimum length of operon. Default is 20.
--statistics, -s Doing statistics for operon analysis. Default is
False. The name of statistics file is -
stat_operon_$STRAIN_NAME.csv.
--combine_gff, -c Convert the operon and all features you assigned to
one gff file. Default is False.
- Output files
All output files will be stored in $ANNOgesic/output/operon
.
gffs
: The gff files which integrate the information of TSSs, annotations,
transcripts, 5’UTRs, and 3’UTRs.
tables
: The tables of operons which store all information of operons and suboperons.
statistics
: Statistics files.
circrna¶
circrna
can detect the potential circular RNAs. It applies Segemehl
to detect circular RNAs. Then it will checks genome annotation files and quality of splicing site detection to
exclude false positive. User can assign reads for mapping and detecting circular RNAs or assign alignment files to skip mapping.
But BE CAREFUL, If user uses alignment files, they should be mapped by Segemehl
and with -S
. Or circrna
can’t find the proper candidates.
- Pre-required tools and files
Fasta files of reads or alignment files. If you want to use alignment files directly, they should be
mapped by Segemehl and with -S
.
Fasta files and gff files of genome.
- Arguments
usage: annogesic circrna [-h] [--segemehl_folder SEGEMEHL_FOLDER]
[--samtools_path SAMTOOLS_PATH] [--align]
[--tex_bam_path TEX_BAM_PATH]
[--fragmented_bam_path FRAGMENTED_BAM_PATH]
[--process PROCESS] [--fasta_path FASTA_PATH]
[--annotation_path ANNOTATION_PATH]
[--support_reads SUPPORT_READS]
[--start_ratio START_RATIO] [--end_ratio END_RATIO]
[--ignore_hypothetical_protein IGNORE_HYPOTHETICAL_PROTEIN]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--segemehl_folder SEGEMEHL_FOLDER, -sg SEGEMEHL_FOLDER
Please assign the folder of segemehl.
--samtools_path SAMTOOLS_PATH, -st SAMTOOLS_PATH
Please assign the path of samtools.
--align, -a Using segemehl to map reads (included splice
detection). If you already usd segemehl with -S to map
your reads, you can skip this step, don't need to turn
it on. Please be attention, it only use default
parameters of segemehl to map your reads. Moreover, it
will map all read files in ANNOgesic/input/reads. If
you want to run some specific functions of segemehl,
please directly run segemehl by yourself. Default is
False.
--tex_bam_path TEX_BAM_PATH, -tb TEX_BAM_PATH
If you already has Bam files, Please assign the TEX+/-
Bam path.
--fragmented_bam_path FRAGMENTED_BAM_PATH, -fb FRAGMENTED_BAM_PATH
If you already has Bam files, Please assign the
fragmented Bam path.
--process PROCESS, -p PROCESS
The number of parallels processes for --align. Default
is 10.
--fasta_path FASTA_PATH, -f FASTA_PATH
The folder of genome fasta.
--annotation_path ANNOTATION_PATH, -g ANNOTATION_PATH
The folder of genome annotation gff files.
--support_reads SUPPORT_READS, -s SUPPORT_READS
The cut off of supported reads. Default is 10.
--start_ratio START_RATIO, -sr START_RATIO
The ratio of (read support circ / all read) at
starting point. The ratio of candidates should higher
than this cutoff. Default is 0.5.
--end_ratio END_RATIO, -er END_RATIO
The ratio of (read support circ / all read) at end
point. The ratio of candidates should higher than this
cutoff. Default is 0.5.
--ignore_hypothetical_protein IGNORE_HYPOTHETICAL_PROTEIN, -ih IGNORE_HYPOTHETICAL_PROTEIN
If you want to ignore hypothetical protein in genome
annotation file, you can turn it on. Default is False.
- Output files
All the output files will be stored in $ANNOgesic/output/circRNA
.
gffs
: Gff files of circular RNAs. $STRAINNAME_best.gff
is the gff files for best result after comparing
with genome annotation and quality of splicing. $STRAINNAME_all.gff
is for all candidates without filering.
circRNA_tables
: The tables for circular RNAs with more details.
statistics
: Statistics files.
segemehl_align
: If circrna
starts from read mapping, the folder is for results of mapping.
segemehl_splice
: The results of splicing detection. The information of the splicing tables, please
refer to Segemehl.
go_term¶
go_term
can compare the genome annotation files and Uniprot to retreive the information of Gene Ontology.
It also provides some analyses of Go terms.
- Pre-required tools and files
idmapping_selected.tab from Uniprot.
Gff files of genome annotation.
If you want to compute the expressed CDSs only, you need to assign the transcript gff files.
- Arguments
usage: annogesic go_term [-h] [--annotation_path ANNOTATION_PATH]
[--transcript_path TRANSCRIPT_PATH]
[--UniProt_id UNIPROT_ID] [--go_obo GO_OBO]
[--goslim_obo GOSLIM_OBO]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--annotation_path ANNOTATION_PATH, -g ANNOTATION_PATH
The path of genome annotation gff folder.
--transcript_path TRANSCRIPT_PATH, -a TRANSCRIPT_PATH
If you assign the path of transcript gff folder, it
can only extract the expressed CDS to retrieve GO
term.
--UniProt_id UNIPROT_ID, -u UNIPROT_ID
The path of UniProt ID mapping database. Default is
ANNOgesic/input/database/idmapping_selected.tab.
--go_obo GO_OBO, -go GO_OBO
The path of go.obo. Default is
ANNOgesic/input/database/go.obo.
--goslim_obo GOSLIM_OBO, -gs GOSLIM_OBO
The path of goslim.obo. Default is
ANNOgesic/input/database/goslim_generic.obo.
- Output files
All output files will be stored in ANNOgesic/output/Go_term
.
Go_term_results
: The tables of Go terms information.
statistics
: Statistics files and figures.
srna_target¶
srna_target
will search the potential targets of sRNAs. User can assign the
programs for running (RNAup or RNAplex or both). We recommand user to run with both
programs. srna_target
can compare the both results and provide the best ones.
- Pre-required tools and files
Gff files of genome annotation.
Gff files of sRNAs.
Fasta files of genome.
- Arguments
usage: annogesic srna_target [-h] [--Vienna_folder VIENNA_FOLDER]
[--annotation_path ANNOTATION_PATH]
[--fasta_path FASTA_PATH] [--sRNA_path SRNA_PATH]
[--query_sRNA QUERY_SRNA] [--program PROGRAM]
[--interaction_length INTERACTION_LENGTH]
[--window_size_target WINDOW_SIZE_TARGET]
[--span_target SPAN_TARGET]
[--window_size_srna WINDOW_SIZE_SRNA]
[--span_srna SPAN_SRNA]
[--unstructured_region_RNAplex_target UNSTRUCTURED_REGION_RNAPLEX_TARGET]
[--unstructured_region_RNAplex_srna UNSTRUCTURED_REGION_RNAPLEX_SRNA]
[--unstructured_region_RNAup UNSTRUCTURED_REGION_RNAUP]
[--energy_threshold ENERGY_THRESHOLD]
[--duplex_distance DUPLEX_DISTANCE] [--top TOP]
[--process_rnaplex PROCESS_RNAPLEX]
[--process_rnaup PROCESS_RNAUP]
[--continue_rnaup]
[--potential_target_start POTENTIAL_TARGET_START]
[--potential_target_end POTENTIAL_TARGET_END]
[--target_feature TARGET_FEATURE]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--Vienna_folder VIENNA_FOLDER
Please assign the folder of Vienna package. It should
include RNAplfold, RNAup and RNAplex.
--annotation_path ANNOTATION_PATH, -g ANNOTATION_PATH
The path of genome annotation gff folder.
--fasta_path FASTA_PATH, -f FASTA_PATH
The path of genome fasta folder.
--sRNA_path SRNA_PATH, -r SRNA_PATH
The path of sRNA gff folder.
--query_sRNA QUERY_SRNA, -q QUERY_SRNA
Please assign the query sRNA. If you want to compute
all sRNA in gff file, please keyin 'all'.the input
format should be like, $STRAIN:$STRAND:$START:$END. If
you want to compute more than one sRNA, please use
comma to separate them.For example,
NC_007795.1:+:200:534,NC_007795.1:-:6767:6900. Default
is all.
--program PROGRAM, -p PROGRAM
Using RNAplex, RNAup or both. Default is both.
--interaction_length INTERACTION_LENGTH, -i INTERACTION_LENGTH
Maximum length of an interaction. Default is 30.
--window_size_target WINDOW_SIZE_TARGET, -wt WINDOW_SIZE_TARGET
Only works when --program is RNAplex or both. Average
the pair probabilities over windows of given size for
RNAplex. Default is 240.
--span_target SPAN_TARGET, -st SPAN_TARGET
Only works when --program is RNAplex or both. Set the
maximum allowed separation of a base pair to span for
RNAplex. Default is 160.
--window_size_srna WINDOW_SIZE_SRNA, -ws WINDOW_SIZE_SRNA
Only works when --program is RNAplex or both. Average
the pair probabilities over windows of given size for
RNAplex. Default is 30.
--span_srna SPAN_SRNA, -ss SPAN_SRNA
Only works when --program is RNAplex or both. Set the
maximum allowed separation of a base pair to span for
RNAplex. Default is 30.
--unstructured_region_RNAplex_target UNSTRUCTURED_REGION_RNAPLEX_TARGET, -ut UNSTRUCTURED_REGION_RNAPLEX_TARGET
Only works when --program is RNAplex or both. Compute
the mean probability that regions of length 1 to a
given length are unpaired for RNAplex. Default is 30.
--unstructured_region_RNAplex_srna UNSTRUCTURED_REGION_RNAPLEX_SRNA, -us UNSTRUCTURED_REGION_RNAPLEX_SRNA
Only works when --program is RNAplex or both. Compute
the mean probability that regions of length 1 to a
given length are unpaired for RNAplex. Default is 30.
--unstructured_region_RNAup UNSTRUCTURED_REGION_RNAUP, -uu UNSTRUCTURED_REGION_RNAUP
Only works when --program is RNAup or both. Compute
the mean probability that regions of length 1 to a
given length are unpaired for RNAplex. Default is 30.
--energy_threshold ENERGY_THRESHOLD, -e ENERGY_THRESHOLD
Only works when --program is RNAplex or both. Minimum
energy for a duplex to be returned for RNAplex.
Default is -8.
--duplex_distance DUPLEX_DISTANCE, -d DUPLEX_DISTANCE
Only works when --program is RNAplex or both. Distance
between target 3' ends of two consecutive duplexes for
RNAplex. Default is 20.
--top TOP, -t TOP The output file only includes --top. Default is 20.
--process_rnaplex PROCESS_RNAPLEX, -pp PROCESS_RNAPLEX
The amount of parallel processes for running RNAplex
prediction. Default is 5.
--process_rnaup PROCESS_RNAUP, -pu PROCESS_RNAUP
The amount of parallel processes for running RNAup
prediction. Default is 20.
--continue_rnaup, -cr
RNAup will take a long time for running if you want to
compute a lot of sRNA. If the process crush, you can
turn it on. This flag will continue running RNAup
based on your previous running. Default is False.
--potential_target_start POTENTIAL_TARGET_START, -ps POTENTIAL_TARGET_START
How many upstream nucleotides of the start point of
CDS that you want to extract as potential target.
Default is 200.
--potential_target_end POTENTIAL_TARGET_END, -pe POTENTIAL_TARGET_END
How many downstream nucleotides of the start point of
CDS that you want to extract as potential target.
Default is 150.
--target_feature TARGET_FEATURE, -tf TARGET_FEATURE
What features you want to extract as potential
targets. If you want to use multi-features, just use
comma to separate them. For example, CDS,gene. Default
is CDS.
- Output files
All output files will be stored in $ANNOgesic/output/sRNA_targets
.
RNAplex
: All results of RNAplex. $STRAIN_RNAplex.txt
is raw results of RNAplex.
It includes the information of binding situations. $STRAIN_RNAplex_rank.csv
is the results
that sorted by binding energy.
RNAup
: All results of RNAup. $STRAIN_RNAup.txt
is raw results of RNAup.
It includes the information of binding situations. $STRAIN_RNAup_rank.csv
is the results
that sorted by binding energy.
merge
: The results which merge RNAplex
and RNAup
. $STRAIN_merge.csv
merges all candidates of both programs.
$STRAIN_overlap.csv
lists the results which be top 20(default) in both methods.
sRNA_seqs
: The fasta sequences of sRNAs.
target_seqs
: The fasta sequences of potential targets.
ppi_network¶
ppi_network
will retrieve the data from STRING.
Then use PIE to search
the literatures to support the protein-protein interaction networks. Therefore,
ppi_network
can generate the protein-protein interaction networks with supported literatures.
- Pre-required tools and files
species.vXXXX.txt from STRING.
Ptt files of genome annotation.
- Arguments
usage: annogesic ppi_network [-h] [--gff_path GFF_PATH]
[--proteinID_strains PROTEINID_STRAINS]
[--without_strain_pubmed]
[--species_STRING SPECIES_STRING] [--score SCORE]
[--node_size NODE_SIZE] [--query QUERY]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--gff_path GFF_PATH, -g GFF_PATH
The path of genome annotation gff folder. Please
confirm your gff files which have proper locus_tag
item in attributes. The locus_tag items can be real
locus tag like locus_tag=SAOUHSC_00003, or they can be
assigned by gene name, such as, locus_tag=dnaA.
--proteinID_strains PROTEINID_STRAINS, -s PROTEINID_STRAINS
This is for query strain and filename. In order to
retrieve the data from STRING and Pubmed, you also
have to assign the similar reference. For example, the
query strain is Staphylococcus aureus HG003, but there
is no Staphylococcus aureus HG003 in STRING database.
However, there is Staphylococcus aureus 8325 which is
highly similar with Staphylococcus aureus HG003.
Therefore, we can use it as reference. you can assign
it like Staphylococcus_aureus_HG003.gff:Staphylococcus
_aureus_HG003:"Staphylococcus aureus
8325":"Staphylococcus aureus". or Staphylococcus_aureu
s_HG003.gff:Staphylococcus_aureus_HG003:"93061":"Staph
ylococcus aureus". or Staphylococcus_aureus_HG003.gff:
Staphylococcus_aureus_HG003:"Staphylococcus aureus
NCTC 8325":"Staphylococcus aureus". (gff_filename:gff_
strain_name:STRING_name:Pubmed_name). First one is the
gff file name. Second one is the strain name of gff
files. Third one is for STRING database, and the
fourth one is for Pubmed. Of course, you can run the
script for several strains at the same time, just put
comma between these strains. Before running it, please
check the species table which should located in
ANNOgesic/input/database .If you didn't download the
file, please download it. You can use taxon_id,
STRING_name_compact or official_name_NCBI to represent
STRING_name.BE CAREFUL, if the name which you assigned
has spaces, please put "" at two ends. For the name of
Pubmed, you can assign the name not so specific. If
you assign a specific name, it may not be able to find
the related literatures.
--without_strain_pubmed, -n
If you want to retrieve pubmed without assign any
strains, please turn it on. Default is False.
--species_STRING SPECIES_STRING, -d SPECIES_STRING
Please assign the path of species table of STRING.
--score SCORE, -ps SCORE
Please assign the cutoff of text-mining score. The
value is from -1 to 1. Default is 0.
--node_size NODE_SIZE, -ns NODE_SIZE
Please assign the size of nodes in figure, default is
4000.
--query QUERY, -q QUERY
Please assign the query protein here. The format is
$STRAINOFGFF:$START_POINT:$END_POINT:$STRAND.If you
want to compute more than one protein, please use
comma to separate them. For example, Staphylococcus_au
reus_HG003:345:456:+.Staphylococcus_aureus_HG003:2000:
3211:-. If you want to compute all protein, just type
all. Default is all.
- Output files
All the output files will be stored in $ANNOgesic/output/PPI
.
best_results
: The results which the score of supported literatures(the text-mining
(PIE) is high enough).
$STRAIN_without_strain.csv
is the results of searching literatures without specific strain.
$STRAIN_with_strain.csv
is the results of searching literatures with specific strain.
For example, Staphylococcus_aureus_8325_without_strain.csv is search with Staphylococcus aureus;
Staphylococcus_aureus_8325_without_strain.csv is search without Staphylococcus aureus.
without_strain
stores all interaction information which search without specific strain.
with_strain
stores all interaction information which search with specific strain.
all_results
: The results of all protein-protein interactions.
(Even the text-mining(PIE) score is too low)
figures
: The thickness represents how many literatures can be found for the interactions.
The solid line means there is some literatures which strongly support the interactions. The dash-dot line
means the supported literatures are very weak. The dot line means there is no literatures which can support the
interactions. The color is the best score of the literatures of the interactions.
subcellular_localization¶
subcellular localization
can predict the subcellular localization of CDSs. It also provide some
statistics and visualization files.
- Pre-required tools and files
Gff files of genome annotation.
Fasta files of genome sequence.
If you want to compute the expressed CDSs only, you need to assign the transcript gff files.
- Arguments
usage: annogesic subcellular_localization [-h] [--Psortb_path PSORTB_PATH]
[--gff_path GFF_PATH]
[--fasta_path FASTA_PATH]
[--transcript_path TRANSCRIPT_PATH]
[--bacteria_type BACTERIA_TYPE]
[--difference_multi DIFFERENCE_MULTI]
[--merge_to_gff]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--Psortb_path PSORTB_PATH
If you want to assign the path of Psortb, please
assign here.
--gff_path GFF_PATH, -g GFF_PATH
The path of genome annotation gff folder.
--fasta_path FASTA_PATH, -f FASTA_PATH
The path of genome fasta folder.
--transcript_path TRANSCRIPT_PATH, -a TRANSCRIPT_PATH
If you assign the path of transcript gff folder, it
can get the results which only include expressed CDS.
--bacteria_type BACTERIA_TYPE, -b BACTERIA_TYPE
Is Gram-positive or Gram-negative. Please assign
'positive' or 'negative'.
--difference_multi DIFFERENCE_MULTI, -d DIFFERENCE_MULTI
If the protein may have multiple locations, it will
calculate the difference of scores(psortb) between
best one and others. If the difference is within this
value, it will also print it out. Default is 0.5. The
maximum value is 10.
--merge_to_gff, -m If you want to merge the information to genome
annotation gff file, please turn it on. Default is
False.
- Output files
All output files will be stored in $ANNOgesic/output/subcellular_localization
.
psortb_results
: The results of Psortb.
statistics
: Statistics files and figures.
riboswitch¶
riboswitch
will search the pre-CDS region which have ribosome binding sites.
Basically, riboswitch
use Infernal to scan
Rfam to find the homologs of known riboswitchs.
- Pre-required tools and files
Rfam.
Gff files of genome annotation.
Fasta files of genome sequence.
File of riboswitch_ID
. The file should contain Rfam ID, Name and Description of riboswitchs.
You can refer to the section of Riboswitch dataset of Rfam
. You can download the file from our
Github (Rfam_riboswitch_ID.csv). You also can create your own one.
- Arguments
usage: annogesic riboswitch [-h] [--infernal_path INFERNAL_PATH]
[--riboswitch_ID RIBOSWITCH_ID]
[--gff_path GFF_PATH] [--tss_path TSS_PATH]
[--UTR_length UTR_LENGTH]
[--transcript_path TRANSCRIPT_PATH]
[--fasta_path FASTA_PATH] [--Rfam RFAM]
[--e_value E_VALUE] [--output_all] [--fuzzy FUZZY]
[--fuzzy_rbs FUZZY_RBS]
[--start_codon START_CODON]
[--max_dist_rbs MAX_DIST_RBS]
[--min_dist_rbs MIN_DIST_RBS]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--infernal_path INFERNAL_PATH, -if INFERNAL_PATH
Please assign the folder of Infernal(where is cmscan
and cmsearch located).
--riboswitch_ID RIBOSWITCH_ID, -i RIBOSWITCH_ID
The path of the riboswitch ID of Rfam. The file is
include the Accession, ID and Description of
riboswitch in Rfam.
--gff_path GFF_PATH, -g GFF_PATH
The path of annotation gff folder.
--tss_path TSS_PATH, -t TSS_PATH
The path of TSS gff folder.
--UTR_length UTR_LENGTH, -u UTR_LENGTH
The length of UTR. Default is 300.
--transcript_path TRANSCRIPT_PATH, -a TRANSCRIPT_PATH
The path of transcript gff folder.
--fasta_path FASTA_PATH, -f FASTA_PATH
The path of genome fasta folder.
--Rfam RFAM, -R RFAM The path of Rfam CM database.
--e_value E_VALUE, -e E_VALUE
The cutoff of e value. Default is 0.001.
--output_all, -o One sequence may fit multiple riboswitches. If you
want to output all of them, please turn it on. Or it
will only print the best one. Default is False.
--fuzzy FUZZY, -z FUZZY
It will extend some nucleotides of 3' or 5' end.
Default is 10.
--fuzzy_rbs FUZZY_RBS, -zr FUZZY_RBS
How many nucleotides of ribosomal binding site allow
to be different with AGGAGG? Default is 2.
--start_codon START_CODON, -ac START_CODON
What kinds of start coden you want to use. If you want
to import more than one type of start codon, please
use comma to separate them. Default is ATG.
--max_dist_rbs MAX_DIST_RBS, -Mr MAX_DIST_RBS
The maximum distance between ribosomal binding site
and start codon. Default is 14.
--min_dist_rbs MIN_DIST_RBS, -mr MIN_DIST_RBS
The minmum distance between ribosomal binding site and
start codon. Default is 5.
- Output files
All output files will be stored in $ANNOgesic/output/riboswitch
.
gffs
: The gff files of riboswitchs.
tables
: The tables of riboswichs with more details.
scan_Rfam
: The results of cmscan
of Infernal.
User can check the results of blast.
statistics
: The statistics files and figures.
optimize_tsspredator¶
optimize_tsspredator
can adapt the parameter set of TSSpredator.
User just need to manual check around 200kb as a reference (we suggest the manual detected TSSs should more than 50).
Then optimize_tsspredator
can scan whole genome based on the principle of manual detection to get the best results.
- Pre-required tools and files
The libraries and wiggle files of TEX +/-.
Please refer to the The format of libraries for import to ANNOgesic
.
Fasta file of genome sequence.
Gff file of genome annotation.
Gff file of manual detection.
- Arguments
usage: annogesic optimize_tsspredator [-h]
[--TSSpredator_path TSSPREDATOR_PATH]
[--fasta_file FASTA_FILE]
[--annotation_file ANNOTATION_FILE]
[--wig_folder WIG_FOLDER]
[--manual MANUAL]
[--strain_name STRAIN_NAME]
[--max_height MAX_HEIGHT]
[--max_height_reduction MAX_HEIGHT_REDUCTION]
[--max_factor MAX_FACTOR]
[--max_factor_reduction MAX_FACTOR_REDUCTION]
[--max_base_height MAX_BASE_HEIGHT]
[--max_enrichment_factor MAX_ENRICHMENT_FACTOR]
[--max_processing_factor MAX_PROCESSING_FACTOR]
[--utr_length UTR_LENGTH] [--lib LIB]
[--output_prefix OUTPUT_PREFIX]
[--cluster CLUSTER] [--length LENGTH]
[--core CORE] [--program PROGRAM]
[--replicate_match REPLICATE_MATCH]
[--steps STEPS]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--TSSpredator_path TSSPREDATOR_PATH
If you want to assign the path of TSSpredator, please
assign here. Default is /usr/local/bin/TSSpredator.jar
--fasta_file FASTA_FILE, -fs FASTA_FILE
Path of one target genome fasta file which you want to
opimize it.
--annotation_file ANNOTATION_FILE, -g ANNOTATION_FILE
Path of one target genome annotation gff file which
you want to opimize it.
--wig_folder WIG_FOLDER, -w WIG_FOLDER
The folder of the TEX+/- wig folder.
--manual MANUAL, -m MANUAL
The file of manual checked gff file.
--strain_name STRAIN_NAME, -n STRAIN_NAME
The name of the strain you want to optimize.
--max_height MAX_HEIGHT, -he MAX_HEIGHT
This value relates to the minimum number of read
starts at a certain genomic position to be considered
as a TSS candidate. During optimization will be never
larger than this value. Default is 2.5.
--max_height_reduction MAX_HEIGHT_REDUCTION, -rh MAX_HEIGHT_REDUCTION
When comparing different strains/conditions and the
step height threshold is reached in at least one
strain/condition, the threshold is reduced for the
other strains/conditions by the value set here. This
value must be smaller than the step height threshold.
During optimization will be never larger than this
value. Default is 2.4.
--max_factor MAX_FACTOR, -fa MAX_FACTOR
This is the minimum factor by which the TSS height has
to exceed the local expression background. During
optimization will be never larger than this value.
Default is 10.
--max_factor_reduction MAX_FACTOR_REDUCTION, -rf MAX_FACTOR_REDUCTION
When comparing different strains/conditions and the
step factor threshold is reached in at least one
strain/condition, the threshold is reduced for the
other strains/conditions by the value set here. This
value must be smaller than the step factor threshold.
During optimization will be never larger than this
value. Default is 9.9.
--max_base_height MAX_BASE_HEIGHT, -bh MAX_BASE_HEIGHT
This is the minimum number of reads should be mapped
on TSS. During optimization will be never larger than
this value. Default is 0.06.
--max_enrichment_factor MAX_ENRICHMENT_FACTOR, -ef MAX_ENRICHMENT_FACTOR
This is the minimum enrichment factor. During
optimization will be never larger than this value.
Default is 6.0.
--max_processing_factor MAX_PROCESSING_FACTOR, -pf MAX_PROCESSING_FACTOR
This is the minimum processing factor. If untreated
library is higher than the treated library and above
which the TSS candidate is considered as a processing
site and not annotated as detected. During
optimization will be never larger than this value.
Default is 6.0
--utr_length UTR_LENGTH, -u UTR_LENGTH
The length of UTR. It is for Primary and Secondary
TSSs. Default is 300.
--lib LIB, -l LIB The libraries of wig files for TSSpredator. The format
is: wig_file_name:tex_treat_or_not(tex or notex):condi
tion_id(integer):replicate_id(alphabet):strand(+ or
-). If you have multiple wig files, please use comma
to separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--output_prefix OUTPUT_PREFIX, -p OUTPUT_PREFIX
The output prefix of all conditions. If you have
multiple conditions, please use comma to separate
them. For example,
prefix_condition1,prefix_condition2.
--cluster CLUSTER, -cu CLUSTER
If the position between manual one and predicted one
is smaller or equal than this value, it will only
print one of them. Default is 2.
--length LENGTH, -le LENGTH
The length of genome for running optimization. If you
don't want to run for whole genome, you can assign the
length of partial genome.Default is compare whole
genome.
--core CORE, -c CORE How many paralle runs do you want to use. Default is
4.
--program PROGRAM, -t PROGRAM
You want to run optimize TSS or processing site.
Please assign "TSS" or "Processint_site". Default is
TSS.
--replicate_match REPLICATE_MATCH, -rm REPLICATE_MATCH
The TSS candidates should match to how many number of
the replicates. Default is 1.
--steps STEPS, -s STEPS
How many steps do you want to run. Default is 4000
runs.
- Output files
Based on the programs (TSS/processing site), the output files will be stored in
$ANNOgesic/output/TSS/optimized_TSSpredator
or $ANNOgesic/output/processing_site/optimized_TSSpredator
.
stat.csv
stores the information of every step. The first column is the number of run.
The second column is the parameter set. he
represents height; rh
represents
height reduction; fa
means factor; rf
means factor reduction; bh
is
base height; ef
is enrichment factor; pf
is processing factor. About the details
of parameters, please refer to TSSpredator.
For example, he_2.0_rh_1.8_fa_4.4_rf_2.8_bh_0.08_ef_3.0_pf_2.6
means height is 2.0,
height reduction is 1.8, factor is 4.4, factor reduction is 2.8, base height is 0.08,
enrichment factor is 3.0 and processing factor is 2.6. The third and fourth columns are
the number of true positive. The fifth and sixth columns are true positive rate. The seventh
and eighth columns are the number of false positive. The ninth and tenth false positive rate.
The eleventh and twelfth columns are the number of false negative. The thirteenth and fourteenth
columns are missing rate.
best.csv
stores the best parameter set. The meanings of all columns are the same as stat.csv
.
screenshot¶
screenshot
will generate batch files for producing screenshot of IGV.
When the batch files produced, user just needs to open IGV, then presses tools
on the top tags and choose run batch script
. The program will automatically produce screenshots.
Then user can refer to them easily.
- Pre-required tools and files
IGV.
Gff files that user want to produce screenshots. All screenshots will be produced based on the positions of main_gff
.
side_gffs
are the gff files that user want to compare with main_gff
. They will also appear in the screenshots.
Fasta files of genome.
The libraries and wiggle files. Please refer to the The format of libraries for import to ANNOgesic
.
- Arguments
usage: annogesic screenshot [-h] [--main_gff MAIN_GFF] [--side_gffs SIDE_GFFS]
[--fasta FASTA]
[--frag_wig_folder FRAG_WIG_FOLDER]
[--tex_wig_folder TEX_WIG_FOLDER]
[--height HEIGHT] [--tex_libs TEX_LIBS]
[--frag_libs FRAG_LIBS] [--present PRESENT]
[--output_folder OUTPUT_FOLDER]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--main_gff MAIN_GFF, -mg MAIN_GFF
Screenshot will based on the position of main_gff file
to generate screenshot.
--side_gffs SIDE_GFFS, -sg SIDE_GFFS
If you want to import some gff files to compare with
--main_gff, please assign here. You can input more
than one gff files, just use comma to separate them.
--fasta FASTA, -f FASTA
The path of genome fasta folder.
--frag_wig_folder FRAG_WIG_FOLDER, -fw FRAG_WIG_FOLDER
If you want to include the information of fragmented
wig file, please assign the folder.
--tex_wig_folder TEX_WIG_FOLDER, -tw TEX_WIG_FOLDER
If you want to include the information of tex+/- wig
file, please assign the folder.
--height HEIGHT, -he HEIGHT
You can assign the height of screenshot. Default is
1500.
--tex_libs TEX_LIBS, -tl TEX_LIBS
If you want to include the tex+/- wig file, please
also assign proper format here. The format is:
wig_file_name:tex_treat_or_not(tex or notex):condition
_id(integer):replicate_id(alphabet):strand(+ or -). If
you have multiple wig files, please use comma to
separate the wig files. For example,
wig1:tex:1:a:+,wig2:tex:1:a:-.
--frag_libs FRAG_LIBS, -fl FRAG_LIBS
If you want to include the fragmented wig file, please
also assign proper format here. The format is: wig_fil
e_name:fragmented(frag):condition_id(integer):replicat
e_id(alphabet):strand(+ or -). If you have multiple
wig files, please use comma to separate the wig files.
For example, wig1:frag:1:a:+,wig2:frag:1:a:-.
--present PRESENT, -p PRESENT
Which type that you want to present in the screenshot.
expand/collapse/squish. Default is expand.
--output_folder OUTPUT_FOLDER, -o OUTPUT_FOLDER
Please assign the output folder. It will create a
sub-folder "screenshots" in the output folder to store
the results.
- Output files
Based on the paths of main_gff
, screenshot
will generate a folder - screenshots
under the
folder of main_gff
. All output files will be stored in this folder.
forward.txt
is the batch file of forward strand for running on IGV.
reverse.txt
is the batch file of reverse strand for running on IGV.
forward
is the folder for storing screenshots of forward strand.
reverse
is the folder for storing screenshots of reverse strand.
When user runs batch files on IGV. The screenshots will automatically store in forward
and reverse
.
The format of filenames will be $STRAIN:$START-$END.png
. For example, NC_007795:1051529-1051696.png
means the strain is NC_007795. The feature’s start point is 1051529 and the end point is
1051696.
color_png¶
color_png
is a following procedure of screenshot
. If there are many wiggle files, it will be difficult
to distinguish the tracks. Especially, when user want to compare TEX +/- libraries to check TSSs or processing sites,
it is not convenient. color_png
can color the tracks based on TEX +/- libraries. Therefore, the user can
refer to the screenshots much easier.
- Pre-required tools and files
The screenshots which generated from screenshot
. Please make sure the folders of forward
and reverse
exist in the folder of screenshots
.
- Arguments
usage: annogesic color_png [-h] [--screenshot_folder SCREENSHOT_FOLDER]
[--track_number TRACK_NUMBER]
[--ImageMagick_covert_path IMAGEMAGICK_COVERT_PATH]
[project_path]
positional arguments:
project_path Path of the project folder. If none is given, the
current directory is used.
optional arguments:
-h, --help show this help message and exit
--screenshot_folder SCREENSHOT_FOLDER, -f SCREENSHOT_FOLDER
The folder which stores the folder of "screenshots"
which generated by subcommand "screenshot".
--track_number TRACK_NUMBER, -t TRACK_NUMBER
How many number of tracks do you have?
--ImageMagick_covert_path IMAGEMAGICK_COVERT_PATH, -m IMAGEMAGICK_COVERT_PATH
Please assign the path of "convert" in ImageMagick
package
- Output files
The new screenshots will replace the previous ones automatically.