Module

- reffa

NGS reference genome usually were FASTA and reffa module mainly focused on FASTA format GENOME file.

ReffaFile were a Python class of FASTA format GENOME file, which can be used to run generate_dict and index by BWA/STAR/Bowtie/Bowtie2.

from iseq.reffa import ReffaFile
from iseq.utils import get_config
config_dict = get_config("/path/config.cfg")
reffa = ReffaFile("/path/hg19.fa", "hg19")
reffa.generate_dict(config_dict)
reffa.bwa_index(config_dict)
reffa.star_index(config_dict)
reffa.bowtie_index(config_dict)
reffa.bowtie2_index(config_dict)

- fastq

NGS sequencing raw data format were FASTQ and fastq module mainly focused on FASTQ format raw sequencing file.

FastqFile were a python class of FASTQ format raw sequencing file, which can be used to run mapping using BWA/STAR/Bowtie/Bowtie2/Tophat/Tophat2

from iseq.fastq import FastqFile
from iseq.utils import get_config
config_dict = get_config("/path/config.cfg")
fq_1 = FastqFile("/path/read_1.fq.gz", sampleid = "example")
fq_2 = "/path/read_2.fq.gz"
fq_1.bwa_mapping(config_dict, "/path/outdir", fq_2)
fq_1.star_mapping(config_dict, "/path/outdir", fq_2)
fq_1.bowtie2_mapping(config_dict, "/path/outdir", fq_2)
fq_1.bowtie_mapping(config_dict, "/path/outdir", fq_2)
fq_1.tophat_mapping(config_dict, "/path/outdir", fq_2)

- sam

NGS sequencing format were SAM after map to FASTA format GENOME without convert to BAM and sam module mainly focused on SAM format sequencing file.

SamFile were a python class of SAM format sequencing file, which can be used to run convert2bam using samtools

from iseq.sam import SamFile
from iseq.utils import get_config
config_dict = get_config("/path/config.cfg")
sam = BamFile("/path/example.sam", sampleid = "example")
sam.convert2bam(config_dict, "/path/outdir/out.bam")

- bam

NGS sequencing format were BAM after map to FASTA format GENOME and bam module mainly focused on BAM format sequencing file.

BamFile were a python class of BAM format sequencing file, which can be used to run index/mpileup using samtools, contig_reorder/add_read_group/mark_duplicates using Picard, realigner_target_creator/indel_realigner/recalibration/print_reads/split_ntrim using GATK for pre-process step, /haplotype_caller/unifiedgenotyper_caller/mutect_caller/varscan_caller/torrent_caller/lofreq_caller/pindel_caller/freebayes_caller for variants discover step.

from iseq.bam import BamFile
from iseq.utils import get_config
config_dict = get_config("/path/config.cfg")
bam = BamFile("/path/example.bam", sampleid = "example")
bam.index(config_dict)
bam.mpileup(config_dict, "/path/outdir/out.mpileup")
bam.contig_reorder(config_dict, "/path/outdir/out.reorder_contig.bam")
...
bam.haplotype_caller(config_dict, "/path/outdir/")
bam.varscan_caller(config_dict, "/path/outdir/")
...
bam.pindel_caller(config_dict, "/path/outdir/")

- vcf

NGS standard Variant Call Format were VCF and vcf module mainly focused on VCF format file.

VcfFile were a python class of VCF format file, which can be used to run fsqd_filter/control_filter/unifiedgenotyper_filter/snpfilter/annovar/merge/varscan2gatkfmt/select_snp/select_indel using GATK and other tools

from iseq.vcf import VcfFile
from iseq.utils import get_config
config_dict = get_config("/path/config.cfg")
case_vcf = VcfFile("/path/example.case.vcf", sampleid = "example")
control_vcf = "/path/example.control.vcf", sampleid = "example"
case_vcf.fsqd_filter(config_dict, "/path/outdir/out.vcf") # GATK VariantFiltration (FS>;QD<)
case_vcf.control_filter(config_dict, control_vcf, "/path/outdir/out.vcf")
case_vcf.snpfilter(config_dict, "/path/outdir/out.vcf") # Filter all position including 'rs' flag
case_vcf.annovar(config_dict, "/path/outdir/") # Using ANNOVAR to annotation variants record
...

- csv / mpileup / result

csv module for ANNOVAR CSV foramt output; mpileup module for samtools mpileup output format; result for user friendly format result file which can be used to publish or report. Which mainly including CsvFile / MpileupFile/ ResultFile class.

• Compute variants depth and variant alle depth.
• Using samtools to mpileup that position
• Others

Pipeline

Without paired normal control sample

# fastq2vcf mode
panel -c config.cfg \
         -s A01A \
         -m fastq2vcf \
         -1 A01A_1.fq.gz \
         -2 A01A_2.fq.gz \
         --bamprocess 00101111 \
         -o outdir

# fastq2bam
panel -c config.cfg \
         -s A01A \
         -m fastq2bam \
         -1 A01A_1.fq.gz \
         -2 A01A_2.fq.gz \
         --bamprocess 00101111 \
         -o outdir

# bam2vcf
panel -c config.cfg \
         -s A01A \
         -m bam2vcf \
         --in_bam A01A.bam \
         --bamprocess 00000000 \
         -o outdir

# genomeindex mode
panel -c config.cfg -m genomeindex

With paired normal control sample

# fastq2vcf mode
panel_somatic -c config.cfg \
                 -s A01 \
                 -m fastq2vcf \
                 -1 A01A_1.fq.gz \
                 -2 A01A_2.fq.gz \
                 -3 A01C_1.fq.gz \
                 -4 A01C_2.fq.gz \
                 --bamprocess 00101111 \
                 -o outdir

# fastq2bam mode
panel_somatic -c config.cfg \
                 -s A01 \
                 -m fastq2bam \
                 -1 A01A_1.fq.gz \
                 -2 A01A_2.fq.gz \
                 -3 A01C_1.fq.gz \
                 -4 A01C_2.fq.gz \
                 --bamprocess 00101111 \
                 -o outdir

# bam2vcf mode
panel_somatic -c config.cfg \
                 -s A01 \
                 -m bam2vcf \
                 --case_in_bam A01A.bam \
                 --control_in_bam A01C.bam \
                 --bamprocess 00000000 \
                 -o outdir

# genomeindex mode
panel_somatic -c config.cfg -m genomeindex