Transcription group entry (5): Sequence Comparison and transcription entry

Transcription group entry (5): Sequence Comparison and transcription entry
Task List

• Comparison Software
• Hisat2 usage
• Download the index file
• Comparison, sorting, and Indexing
• Quality Control
• Load IGV, several genes

Hisat2 is compared to the genome, so gapped or splices er is used. This process has been updated. TopHat was first published seven years ago. The comparison speed of STAR is 50 times that of TopHat, and HISAT is 1.2 times that of STAR. HISAT2 is the successor of TopHat2/Bowti2. It uses the improved BWT algorithm to achieve faster speed and less resource occupation. The authors recommend TopHat2/Bowti2 and HISAT users to switch to HISAT2. Official Website: https://ccb.jhu.edu/software/hisat2/index.shtml (the best way to learn a software is to combine the existing Chinese information, coupled with reading the official documentation and HELP documentation, generally at the beginning of learning to use the default parameters, do not adjust parameters) download the index file

cd ~/referencemkdir -p index/hisat && cd index/hisatwget -c ftp://ftp.ccb.jhu.edu/pub/infphilo/hisat2/data/hg19.tar.gzwget -c ftp://ftp.ccb.jhu.edu/pub/infphilo/hisat2/data/mm10.tar.gztar zxvf hg19.tar.gztar xvzf mm10.tar.gz

-C: Use resumable upload to compare, sort, and index the fastq-format reads to obtain the sam file, and then use samtools to convert it to the bam file, and sorting (note the differences between N and P) (you can use pipelines to save the process of saving the intermediate SAM and directly output the BAM file). Write the bash Script: map. sh

#! usr/bin/bashset -uset -eset -o pipefailhg19_ref=/mnt/hgfs/2017/reference/index/hisat/hg19/genomemm10_ref=/mnt/hgfs/2017/reference/index/hisat/mm10/genomedata_path=/mnt/hgfs/2017/rna_seq/dataNUM_THREADS=25ls --color=never Homo*1.fastq.gz | while read id;do(~/biosoft/hisat2-2.1.0/hisat2 -t -p $NUM_THREADS -x $hg19_ref -1 $data_path/${id%_*}_1.fastq.gz -2 $data_path/${id%_*}_2.fastq.gz 2 > ${id%_*}_map.log | samtools view -Sb  - > ${id%_*}.bam);donels --color=never Mus*1.fastq.gz | while read id;do(~/biosoft/hisat2-2.1.0/hisat2 -t -p $NUM_THREADS -x $mm10_ref -1 $data_path/${id%_*}_1.fastq.gz -2 $data_path/${id%_*}_2.fastq.gz 2 > ${id%_*}_map.log | samtools view -Sb  - > ${id%_*}.bam);donels --color=never *.bam | while read id;do(samtools sort --threads $NUM_THREADS $id -o ${id%.*}_sorted.bam);donels --color=never *_sorted.bam | while read id;do(samtools index $id);done

Run the script:

bash map.sh

Quality control the percentage of reads in comparing the quality control (Evaluation and comparison of quality indicators) after comparing the bam files with simple QCReads; check whether the coverage of Reads is consistent with that of the reference chain. Compare to the genome sequence and multi-ratio reads. In addition to Picard, RSeQC, and Qualimap, there are a lot of quality control software.