Ns126:Calendar/NOTES/2016-1-5: Difference between revisions
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*RRBS Analysis Protocol: [[File:RRBS Guide-Baraham.pdf]] | *RRBS Analysis Protocol: [[File:RRBS Guide-Baraham.pdf]] | ||
Normal tissues from RRBS data were downloaded from the ENCODE USCS genome browser (http://hgdownload.cse.ucsc.edu/goldenPath/hg19/encodeDCC/wgEncodeHaibMethylRrbs/). Raw sequence data from 16 tissues were collected (Supplementary Table). Sequences were mapped to the bisulfite-converted human genome (NCBI hg19) using Bismark [1], after trimming the first three nucleotides, using the following parameters: --bowtie2 --phred64-quals --fastq -L 20 -N 1. | *Normal tissues from RRBS data were downloaded from the ENCODE USCS genome browser (http://hgdownload.cse.ucsc.edu/goldenPath/hg19/encodeDCC/wgEncodeHaibMethylRrbs/). Raw sequence data from 16 tissues were collected (Supplementary Table). Sequences were mapped to the bisulfite-converted human genome (NCBI hg19) using Bismark [1], after trimming the first three nucleotides, using the following parameters: --bowtie2 --phred64-quals --fastq -L 20 -N 1. | ||
1. Krueger, F. and S.R. Andrews, Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics, 2011. 27(11): p. 1571-2. | 1. Krueger, F. and S.R. Andrews, Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics, 2011. 27(11): p. 1571-2. |
Revision as of 18:51, 11 January 2016
Collection RRBS Dataset From Richard Myers, HAIB (Encode Project)
Data Description
- RRBS Protocol: File:Myers Lab RRBS Protocol 6-18-2010.pdf
- RRBS Analysis Protocol: File:RRBS Guide-Baraham.pdf
- Normal tissues from RRBS data were downloaded from the ENCODE USCS genome browser (http://hgdownload.cse.ucsc.edu/goldenPath/hg19/encodeDCC/wgEncodeHaibMethylRrbs/). Raw sequence data from 16 tissues were collected (Supplementary Table). Sequences were mapped to the bisulfite-converted human genome (NCBI hg19) using Bismark [1], after trimming the first three nucleotides, using the following parameters: --bowtie2 --phred64-quals --fastq -L 20 -N 1.
1. Krueger, F. and S.R. Andrews, Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics, 2011. 27(11): p. 1571-2.
Fastq Download
- Fastq donwload address: File:Haib.download.files.txt
xargs -n 1 curl -O -L < haib.download.files.txt
Check Phred Score
perl ~/bin/checkphred.pl *fastq
Fastq Quality Control
# trim_galore --phred64 --fastqc --illumina --non_directional --rrbs *.fastq trim_galore --phred64 --fastqc --illumina --rrbs *.fastq
Fastq Alignment
bismark --bowtie2 --phred64-quals --fastq -L 30 -N 1 /home/shg047/db/aligndb/hg19/bismark -1 ENCFF000MLM_trimmed.fq -o ../bam
Methylation haploinfo
bismark_methylation_extractor \ --single-end \ --bedGraph \ --buffer_size 2G \ --remove_spaces \ --zero_based \ --merge_non_CpG \ --comprehensive \ --output ../methyfreq \ ENCFF000MLM_trimmed.fq_bismark_bt2.bam
bismark_methylation_extractor --single-end --bedGraph --buffer_size 2G --remove_spaces --zero_based --merge_non_CpG --comprehensive --output ../methyfreq ENCFF000MLM_trimmed.fq_bismark_bt2.bam
Methylfreq Data Output
- Bismark Run Perl Beginning Script
#/usr/bin/perl use strict; use Cwd; my $dir=getcwd; chdir $dir; my @file=glob("*.fastq"); foreach my $file(@file){ open OUT,">$file.bismark.sh"; print OUT "cd $dir\n"; chomp(my $phredcheck=`perl /home/shg047/bin/checkphred.pl $file`); my ($phred)=split /\s+/,$phredcheck; my $phred="--phred$phred"; print OUT "trim_galore $phred --fastqc --illumina --non_directional --rrbs $file\n"; print OUT "bismark --bowtie2 --phred64-quals --fastq -L 30 -N 1 /home/shg047/db/aligndb/hg19/bismark -1 $file\_qual_trimmed.fastq -o ../bam\n"; print OUT "bismark_methylation_extractor --single-end --bedGraph --buffer_size 2G --remove_spaces --zero_based --merge_non_CpG --comprehensive -- output ../me }
- Bismark Alignment Information Collection Script
- Mapping efficency: File:BismarkAlignment.txt