本次使用的是自己测序所得的cleandata
1.数据去除接头
#使用的trim_galore 过滤
trim_galore -q 20 --phred33 --stringency 3 --length 20 -e 0.1 --paired ./RNAm6Ainput2-7_FKDL220023876-1a_1.clean.fq.gz ./RNAm6Ainput2-7_FKDL220023876-1a_2.clean.fq.gz -o ./
trim_galore -q 20 --phred33 --stringency 3 --length 20 -e 0.1 --paired ./RNAm6AIP2-7_FKDL220023877-1a_1.clean.fq.gz ./RNAm6AIP2-7_FKDL220023877-1a_2.clean.fq.gz -o ./
2.去除adaptor以后进行数据质控
fastqc -t 8 -o /public/home/RNAIP/RNAm6AIP2-7_FKDL220023877-1a RNAm6AIP2-7_FKDL220023877-1a_1.clean_val_1.fq.gz \
fastqc -t 8 -o /public/home/RNAIP/RNAm6AIP2-7_FKDL220023877-1a RNAm6AIP2-7_FKDL220023877-1a_2.clean_val_2.fq.gz
fastqc -t 8 -o /public/home/RNAIP/RNAm6Ainput2-7_FKDL220023876-1a RNAm6Ainput2-7_FKDL220023876-1a_1.clean_val_1.fq.gz \
fastqc -t 8 -o /public/home/RNAIP/RNAm6Ainput2-7_FKDL220023876-1a RNAm6Ainput2-7_FKDL220023876-1a_2.clean_val_2.fq.gz
3.使用bowtie 比对
bowtie2 -x /public/home/genome/bowtie_index/NIP -1 /public/home/RNAIP/RNAm6AIP2-7_FKDL220023877-1a/RNAm6AIP2-7_FKDL220023877-1a_1.clean_val_1.fq.gz -2 /public/home/RNAIP/RNAm6AIP2-7_FKDL220023877-1a/RNAm6AIP2-7_FKDL220023877-1a_2.clean_val_2.fq.gz |/public/anaconda2/envs/chipseq/bin/samtools sort -@ 8 -O bam -o /public/RNAIP_1.bam
bowtie2 -x /public/home/genome/bowtie_index/NIP -1 /public/home/RNAIP/RNAm6Ainput2-7_FKDL220023876-1a/RNAm6Ainput2-7_FKDL220023876-1a_1.clean_val_1.fq.gz -2 /public/home/RNAIP/RNAm6Ainput2-7_FKDL220023876-1a/RNAm6Ainput2-7_FKDL220023876-1a_2.clean_val_2.fq.gz |/public/home/anaconda2/envs/chipseq/bin/samtools sort -@ 8 -O bam -o /public/RNAinput-1.bam
4.选用sambamba对来去重复
sambamba markdup -r RNAinput-1.bam RNAinput.sambamba.rmdup.bam
sambamba markdup -r RNAIP_1.bam RNAIP.sambamba.rmdup.bam
5.smart tool对比对结果排序
samtools sort -O bam -@ 4 -o ./RNAinput.fraw.bam RNAinput.sambamba.rmdup.bam
samtools sort -O bam -@ 4 -o ./RNAIP.fraw.bam RNAIP.sambamba.rmdup.bam
6.smart tool提取可靠的比对结果
samtools view -f 2 -q 30 -o RNAinput.fraw.q30.bam RNAinput.fraw.bam
samtools view -f 2 -q 30 -o RNAIP.fraw.q30.bam RNAIP.fraw.bam
7.call peaks
macs2 callpeak -f BAMPE -c RNAinput.fraw.q30.bam -t RNAIP.fraw.q30.bam -p 0.05 -g 380699722 -n 6mAIP --outdir ./peak --nomodel -B --SPMR
