！RNAseq:上游

2022-02-23 本文已影响0人一只小脑斧

fastp：质控、过滤、去接头（新集群base）

####安装

conda install -c bioconda fastp

conda install -c bioconda/label/cf201901 fastp

#######查看MD5

md5sum

#fastp参数说明：https://www.jianshu.com/p/8fa2ed15dfaa

for i in {4..9} ;

fastp -i LR22A21DX12${i}_Clean_Data1.fq.gz -o LR22A21DX12${i}_1.fq.gz -I LR22A21DX12${i}_Clean_Data2.fq.gz -O LR22A21DX12${i}_2.fq.gz \

done

fastqc质控(新集群base)

fastqc -t 10 *.fq.gz -o /home/yifan/project/XCL/22.2.RNA/Data/CleanData/qc.results

multiqc汇总质控(新集群base)

#安装

conda install -c bioconda multiqc

conda install -c bioconda/label/cf201901 multiqc

#运行

multiqc *_fastqc.zip

STAR比对 RSEM定量(用fastp之后的cleandata)

#下载软件

#STAR conda安装

conda install -c bioconda star

conda install -c bioconda/label/cf201901 star

#STAR 手动安装

Release STAR 2.7.3a ______ 2019/10/08 · alexdobin/STAR · GitHub

tar -xzf STAR_2.4.2a.tar.gz

cd STAR_2.4.2a

# Build STAR

cd source

make STAR

#RSEM

conda install -c bioconda rsem

conda install -c bioconda/label/cf201901 rsem

RNA-Seq gene expression estimation with read mapping uncertainty (deweylab.github.io)

##########构建索引

https://www.plob.org/article/22918.html 里面的bowtie2换成star就行

(104条消息) 使用RSEM进行转录组测序的差异表达分析_TOP生物信息-CSDN博客

#1.人的

~/biosoft/rsem/RSEM-1.3.0/rsem-prepare-reference-gtf ~/annotation/hg38/gencode.v27.annotation.gtf --star~/reference/genome/hg38/GRCh38.p10.genome.fa ~/reference/index/RSEM/hg38/hg38

#2.鼠的

touch rsem.star.ref.new.sh

#PBS -N rsem.star.ref

#PBS -o /home/yifan/data/ref/rsem.star.GRCm38/my.out

#PBS -e /home/yifan/data/ref/rsem.star.GRCm38/my.err

#PBS –l nodes=2:ppn=8

cd /home/yifan/software/RSEM-1.3.3

rsem-prepare-reference --gtf /home/yifan/data/ref/mouse/Mus_musculus.GRCm38.90.gtf \

-p 8 \

--star \

/home/yifan/data/ref/GRCm38/Mus_Mus_musculus.GRCm38.dna.primary_assembly.fa\

/home/yifan/data/ref/rsem.star.GRCm38/GRCm3

#投递任务

qsub rsem.star.ref.new.sh

65691.mu01

####运行

25 26 27 28 29

#PBS -N rsem.26

#PBS -o /home/yifan/project/XCL/22.2.RNA/Data/CleanData/fastp.results/my.out.26

#PBS -e /home/yifan/project/XCL/22.2.RNA/Data/CleanData/fastp.results/my.err.26

#PBS –l nodes=2:ppn=6

outdir=/home/yifan/project/XCL/22.2.RNA/Data/CleanData/rsemresults

index=/home/yifan/data/ref/rsem.star.GRCm38

cd /home/yifan/project/XCL/22.2.RNA/Data/CleanData/fastp.results

for id in 24

fastq1=LR22A21DX1${id}_1.fq.gz

fastq2=LR22A21DX1${id}_2.fq.gz

rsem-calculate-expression -p 10 --no-bam-output --star --star-gzipped-read-file --star-output-genome-bam --estimate-rspd --time --paired-end $fastq1 $fastq2 $index/GRCm38 $outdir/${id}