NGS_RNA-Seq

@NGS_RNA-Seq

NGS_RNA-Seq
####method follow HPC ARS https://bioinformaticsworkbook.org/dataAnalysis/RNA-Seq/RNA-SeqIntro/RNAseq-using-a-genome.html

#####产生以ht2结尾的基因自索引文件
[jinlong.liu@ceres ~]$ cd /project/soy_dis_assembly/NGS_JINLONG/arabidopsis_gene_family/ncbi-genomes-2020-05-09
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ module load hisat2
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ hisat2-build ./GCF_000001735.4_TAIR10.1_genomic.fna GCF_000001735.4_TAIR10.1_genomic

[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ ll

total 296037
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 44026764 May 9 00:48 GCF_000001735.4_TAIR10.1_genomic.1.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 29870612 May 9 00:48 GCF_000001735.4_TAIR10.1_genomic.2.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 3257 May 9 00:47 GCF_000001735.4_TAIR10.1_genomic.3.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 29870607 May 9 00:47 GCF_000001735.4_TAIR10.1_genomic.4.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 52471411 May 9 00:48 GCF_000001735.4_TAIR10.1_genomic.5.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 30416712 May 9 00:48 GCF_000001735.4_TAIR10.1_genomic.6.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 8 May 9 00:47 GCF_000001735.4_TAIR10.1_genomic.7.ht2
-rw-rw-r–. 1 jinlong.liu proj-soy_dis_assembly 8 May 9 00:47 GCF_000001735.4_TAIR10.1_genomic.8.ht2
-rw-r–r--. 1 jinlong.liu proj-soy_dis_assembly 121164916 May 9 00:08 GCF_000001735.4_TAIR10.1_genomic.fna

###利用测序原始数据fastq文件产生比对文件sam
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ module purge
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ module load hisat2
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ module load samtools
[jinlong.liu@ceres ncbi-genomes-2020-05-09]$ module list
Currently Loaded Modules:

1. hisat2/2.0.5 2) curl/7.54.0 3) samtools/1.9
   [jinlong.liu@ceres ncbi-genomes-2020-05-09]$ hisat2 -p 8 -x ./GCF_000001735.4_TAIR10.1_genomic -1 /project/soy_dis_assembly/NGS_JINLONG/arabidopsis_gene_family/SRR4420293_1.fastq -2 /project/soy_dis_assembly/NGS_JINLONG/arabidopsis_gene_family/SRR4420293_2.fastq -S SRR4420293_new.sam

16473386 reads; of these:
16473386 (100.00%) were paired; of these:
1176268 (7.14%) aligned concordantly 0 times
5764718 (34.99%) aligned concordantly exactly 1 time
9532400 (57.87%) aligned concordantly >1 times
----
1176268 pairs aligned concordantly 0 times; of these:
36131 (3.07%) aligned discordantly 1 time
----
1140137 pairs aligned 0 times concordantly or discordantly; of these:
2280274 mates make up the pairs; of these:
1233959 (54.11%) aligned 0 times
261416 (11.46%) aligned exactly 1 time
784899 (34.42%) aligned >1 times
96.25% overall alignment rate

[jinlong.l