Difference between revisions of "1000 Genomes Project Pilot 1 SNP Calling"
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(file size) BAM (binary SAM) file: up to 43Gb (average 10s Gb) for each individual<br>(file size) GLF (binary) file: up to 19Gb (average 10s Gb) for each individual | (file size) BAM (binary SAM) file: up to 43Gb (average 10s Gb) for each individual<br>(file size) GLF (binary) file: up to 19Gb (average 10s Gb) for each individual | ||
| − | + | == (2) Split GLF files by chromosome == | |
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/home1/ylwtx/2009.08.GLF-split/ | /home1/ylwtx/2009.08.GLF-split/ | ||
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Tom suggested combing the first two steps using the following samtools command:<br>samtools -view -u *.bam 22 | samtools pileup –g - > *.glf | Tom suggested combing the first two steps using the following samtools command:<br>samtools -view -u *.bam 22 | samtools pileup –g - > *.glf | ||
| − | + | == (3) Build a list of individuals within each population == | |
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all | ||
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Note: Check to make sure that all the individuals with GLF are included in the list “NA.number.by.popn” | Note: Check to make sure that all the individuals with GLF are included in the list “NA.number.by.popn” | ||
| − | + | == (4) Link files and tabulate # of files per population, per platform == | |
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/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all | ||
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key command: ln -s | key command: ln -s | ||
| − | + | == (5) Check total (aggregated over all individual samples) depth for each population, each platform == | |
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/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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Other: total depth for each site (for vcf) (run after filter)<br> IMPORTANT/total_depth_per_site.r3.csh<br> key command : DepthPerSite<br> Source codes : /home/ylwtx/codes/cpp/ DepthPerSite/<br> Input : GLF files<br> Output : total depth for each site | Other: total depth for each site (for vcf) (run after filter)<br> IMPORTANT/total_depth_per_site.r3.csh<br> key command : DepthPerSite<br> Source codes : /home/ylwtx/codes/cpp/ DepthPerSite/<br> Input : GLF files<br> Output : total depth for each site | ||
| − | + | == (6) Filter sites with total depth at the extremes, within each population, each platform == | |
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/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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key command : glfMerge or glfMerge_noBGZF<br>Source codes : /home/ylwtx/codes/cpp/glfMerge<br> Originally ~goncalo/code/glfMerge<br>Input : GLF files for one individual<br>Output : GLF file | key command : glfMerge or glfMerge_noBGZF<br>Source codes : /home/ylwtx/codes/cpp/glfMerge<br> Originally ~goncalo/code/glfMerge<br>Input : GLF files for one individual<br>Output : GLF file | ||
| − | + | == (8) Promote a set of sites for each population == | |
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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Parameters set:<br>(1) Posterior probability (for being a polymorphism) threshold: 0.999 (0.9 for genomewide but need test)<br>(2) minMapQ = 30<br>(3) –allhet default is ON<br>Input: GLF<br>Output: simplified GLF with three likelihoods | Parameters set:<br>(1) Posterior probability (for being a polymorphism) threshold: 0.999 (0.9 for genomewide but need test)<br>(2) minMapQ = 30<br>(3) –allhet default is ON<br>Input: GLF<br>Output: simplified GLF with three likelihoods | ||
| − | + | == (9) Merge with genotype data == | |
| − | ( | + | === (9.1) prepare genotype data in a unified format === |
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/genotypes_all_2 | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/genotypes_all_2 | ||
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*special thanks to Wei Chen for preparing the genotype files | *special thanks to Wei Chen for preparing the genotype files | ||
| − | ( | + | === (9.2) merge genotype data with sequence data at promoted sites === |
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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Notes: <br> Sites with more than two alleles (not including REF_ALLELE) will be discarded | Notes: <br> Sites with more than two alleles (not including REF_ALLELE) will be discarded | ||
| − | + | == (10) Run thunder (hidden Markov model) == | |
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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Notes:<br>(1) Cleaned monomorphic sites before feeding to thunder (no need, b/c thunder 005 handles AL1/-)<br>(2) All sites are bi-allelic with one of the alleles being the reference allele (sites with more than 2 alleles including the reference allele are discarded at the beginning of thunder run: initially because of a prior dependent on the reference allele. In the current setting, where Freq1 is used for the prior, we can choose to ignore the reference allele information.) <br>a. Codes changed on 2009-11-02<br>(3) Split: | Notes:<br>(1) Cleaned monomorphic sites before feeding to thunder (no need, b/c thunder 005 handles AL1/-)<br>(2) All sites are bi-allelic with one of the alleles being the reference allele (sites with more than 2 alleles including the reference allele are discarded at the beginning of thunder run: initially because of a prior dependent on the reference allele. In the current setting, where Freq1 is used for the prior, we can choose to ignore the reference allele information.) <br>a. Codes changed on 2009-11-02<br>(3) Split: | ||
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Total 150 jobs (50 jobs for each population) | Total 150 jobs (50 jobs for each population) | ||
| − | + | == (11) Ligate thunder results for larger chromosomes == | |
<br>/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all | <br>/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all | ||
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ligate.all. 2009-10-27.csh | ligate.all. 2009-10-27.csh | ||
| − | + | == (12) Extract QC+ sites == | |
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/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/ | ||
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(1) Check for individuals who are genotyped only:<br>For example, before 2009-12, in CHB+JPT, the following 2 individuals are sequenced:<br>NA18631<br>NA18634<br>(2) Check for trios whose sequencing information were not used: e.g., daughter and father of the trio | (1) Check for individuals who are genotyped only:<br>For example, before 2009-12, in CHB+JPT, the following 2 individuals are sequenced:<br>NA18631<br>NA18634<br>(2) Check for trios whose sequencing information were not used: e.g., daughter and father of the trio | ||
| − | + | == (13) Generate other information for VCF format (no longer needed, already generated) == | |
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/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/<br>site.depth.csh (no longer needed, generated in GLF step) | /home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/<br>site.depth.csh (no longer needed, generated in GLF step) | ||
| − | + | == (14) Generate VCF == | |
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/home/ylwtx/1000Genomes/UoM_2009_12<br>cmd.csh<br> $pop.sh<br> vcf.py | /home/ylwtx/1000Genomes/UoM_2009_12<br>cmd.csh<br> $pop.sh<br> vcf.py | ||
| − | + | == (15) Quality check == | |
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~ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all/chr20/tout/CEU | ~ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all/chr20/tout/CEU | ||
| − | (A) Accuracy of genotype calls<br>eval.Rsq.csh<br>or eval.r2_hat.csh<br>(B) Accuracy of haplotype calls<br>comparehaplotypes.csh | + | (A) Accuracy of genotype calls<br>eval.Rsq.csh<br>or eval.r2_hat.csh<br>(B) Accuracy of haplotype calls<br>comparehaplotypes.csh |
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Revision as of 02:40, 17 February 2010
Michigan Pilot One SNP calling work flow:
(1) Create GLF files from BAM files
(2) Split GLF files by chromosome
(3) Build a list of individuals within each population
(4) Link files and tabulate # of files per population, per platform
(5) Check total depth for each population, each platform
(6) Filter sites with total depth at the extremes, within each population, each platform
(7) Obtain one merged GLF for each individual by merging GLFs across platforms for the same individual
(8) Promote a set of sites for each population
(9) Merge with genotype data
(10) Run thunder
(11) Ligate thunder results for larger chromosomes
(12) Extract QC+ sites
(13) Generate other information for VCF format
(14) Generate VCF
(15) Quality check
(1) Create GLF files from BAM files
/home5/2009.08.GLF-1/ update-glf.csh
sam2glf.csh
key command: samtools pileup
Source codes: Check with Paul for the latest version
output GLF format: bgzf
(file size) BAM (binary SAM) file: up to 43Gb (average 10s Gb) for each individual
(file size) GLF (binary) file: up to 19Gb (average 10s Gb) for each individual
(2) Split GLF files by chromosome
/home1/ylwtx/2009.08.GLF-split/
update-glf.csh
splitGLF.csh
key command: glfSplit
Source codes: ~goncalo/code/glfSplit/
Input GLF format: gz or bgzf
Output GLF format: gz
Tom suggested combing the first two steps using the following samtools command:
samtools -view -u *.bam 22 | samtools pileup –g - > *.glf
(3) Build a list of individuals within each population
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all
STEP 0 in s1-5.csh
Note: Check to make sure that all the individuals with GLF are included in the list “NA.number.by.popn”
(4) Link files and tabulate # of files per population, per platform
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all
STEP 1 in s1-5.csh
key command: ln -s
(5) Check total (aggregated over all individual samples) depth for each population, each platform
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 2 in s1-5.csh
key command : DepthDistr
Source codes : /home/ylwtx/codes/cpp/DepthDistr/
Input : GLF files
Output : PMF and CDF of total depth distribution
Other: total depth for each site (for vcf) (run after filter)
IMPORTANT/total_depth_per_site.r3.csh
key command : DepthPerSite
Source codes : /home/ylwtx/codes/cpp/ DepthPerSite/
Input : GLF files
Output : total depth for each site
(6) Filter sites with total depth at the extremes, within each population, each platform
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 3 in s1-5.csh
key command : filterGLF or filterGLF_notBGZF
Source codes : /home/ylwtx/codes/cpp/filterGLF
Input : GLF files
Output : GLF files
Parameters:
minMapQ = 30
back
(7) Obtain one merged GLF for each individual by merging GLFs across platforms for the same individual
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 4 in s1-5.csh
key command : glfMerge or glfMerge_noBGZF
Source codes : /home/ylwtx/codes/cpp/glfMerge
Originally ~goncalo/code/glfMerge
Input : GLF files for one individual
Output : GLF file
(8) Promote a set of sites for each population
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 5 in s1-5.csh
key command : GPT_Freq
Source codes : /home/ylwtx/codes/cpp/glfmultipes_prethunder/GPT_Freq_old
Originally ~goncalo/code/glfMultiples
Parameters set:
(1) Posterior probability (for being a polymorphism) threshold: 0.999 (0.9 for genomewide but need test)
(2) minMapQ = 30
(3) –allhet default is ON
Input: GLF
Output: simplified GLF with three likelihoods
(9) Merge with genotype data
(9.1) prepare genotype data in a unified format
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/genotypes_all_2
See: readme.procedure.txt*
key command : ped_prethunder_freq
Source codes : /home/ylwtx/codes/cpp/ped_prethunder
- special thanks to Wei Chen for preparing the genotype files
(9.2) merge genotype data with sequence data at promoted sites
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 6 in s1-5.csh
key command : merge_prethunder_freq
Source codes : /home/ylwtx/codes/cpp/merge_prethunder
Notes:
Sites with more than two alleles (not including REF_ALLELE) will be discarded
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
STEP 7 in s1-5.csh
Or prep_and_get_thunder_jobs.chr.*csh
key command : thunder_glf_freq
Source codes : /home/ylwtx/codes/cpp/mach-1.0.16
Notes:
(1) Cleaned monomorphic sites before feeding to thunder (no need, b/c thunder 005 handles AL1/-)
(2) All sites are bi-allelic with one of the alleles being the reference allele (sites with more than 2 alleles including the reference allele are discarded at the beginning of thunder run: initially because of a prior dependent on the reference allele. In the current setting, where Freq1 is used for the prior, we can choose to ignore the reference allele information.)
a. Codes changed on 2009-11-02
(3) Split:
Total 150 jobs (50 jobs for each population)
(11) Ligate thunder results for larger chromosomes
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all
ligate.all. 2009-10-27.csh
(12) Extract QC+ sites
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
share.QC+hap. csh (moved to share/ but works under the parent dir, with and without QC)
share/share.csh (with and without QC)
share.QC+dose.csh (moved to share/ but works under the parent dir, with and without QC)
without QC: for VCF
share/calcLD.csh
share/runBatches.LD.csh
share/ batch.LD.pl
share/noSingleton.CEU.csh
share/noSingleton.CHB+JPT.csh
share/noSingleton.YRI.csh
(1) Check for individuals who are genotyped only:
For example, before 2009-12, in CHB+JPT, the following 2 individuals are sequenced:
NA18631
NA18634
(2) Check for trios whose sequencing information were not used: e.g., daughter and father of the trio
(13) Generate other information for VCF format (no longer needed, already generated)
/home/ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.09.all/
site.depth.csh (no longer needed, generated in GLF step)
(14) Generate VCF
/home/ylwtx/1000Genomes/UoM_2009_12
cmd.csh
$pop.sh
vcf.py
(15) Quality check
~ylwtx/codes/cpp/mach-1.0.16/test_thunder/2009.11.all/chr20/tout/CEU
(A) Accuracy of genotype calls
eval.Rsq.csh
or eval.r2_hat.csh
(B) Accuracy of haplotype calls
comparehaplotypes.csh
