reads比对

项目地址：https://github.com/lh3/minimap2。和blast类似，也是局部比对。环状参考序列可以考虑double，解决首尾问题。

# 安装
conda create -n minimap2 -c conda-forge -c bioconda minimap2
conda activate minimap2

# 主要参数
-t：线程数
-a：输出sam，默认paf

# long sequences against a reference genome
./minimap2 -a test/MT-human.fa test/MT-orang.fa > test.sam
# create an index first and then map
./minimap2 -x map-ont -d MT-human-ont.mmi test/MT-human.fa
./minimap2 -a MT-human-ont.mmi test/MT-orang.fa > test.sam
# use presets (no test data)
./minimap2 -ax map-pb ref.fa pacbio.fq.gz > aln.sam       # PacBio CLR genomic reads
./minimap2 -ax map-ont ref.fa ont.fq.gz > aln.sam         # Oxford Nanopore genomic reads
./minimap2 -ax map-hifi ref.fa pacbio-ccs.fq.gz > aln.sam # PacBio HiFi/CCS genomic reads (v2.19 or later)
./minimap2 -ax asm20 ref.fa pacbio-ccs.fq.gz > aln.sam    # PacBio HiFi/CCS genomic reads (v2.18 or earlier)
./minimap2 -ax sr ref.fa read1.fa read2.fa > aln.sam      # short genomic paired-end reads
./minimap2 -ax splice ref.fa rna-reads.fa > aln.sam       # spliced long reads (strand unknown)
./minimap2 -ax splice -uf -k14 ref.fa reads.fa > aln.sam  # noisy Nanopore Direct RNA-seq
./minimap2 -ax splice:hq -uf ref.fa query.fa > aln.sam    # Final PacBio Iso-seq or traditional cDNA，-uf只考虑正义链转录本；-C5对于识别不同物种的剪切位点，灵敏度更高；-O6,24 -B4可以找到更多外显子
./minimap2 -ax splice --junc-bed anno.bed12 ref.fa query.fa > aln.sam  # prioritize on annotated junctions
./minimap2 -cx asm5 asm1.fa asm2.fa > aln.paf             # intra-species asm-to-asm alignment
./minimap2 -x ava-pb reads.fa reads.fa > overlaps.paf     # PacBio read overlap
./minimap2 -x ava-ont reads.fa reads.fa > overlaps.paf    # Nanopore read overlap
# man page for detailed command line options
man ./minimap2.1

Usage: paftools.js <command> [arguments]
Commands:
  view       convert PAF to BLAST-like (for eyeballing) or MAF
  splice2bed convert spliced alignment in PAF/SAM to BED12
  sam2paf    convert SAM to PAF
  delta2paf  convert MUMmer's delta to PAF
  gff2bed    convert GTF/GFF3 to BED12

  stat       collect basic mapping information in PAF/SAM
  liftover   simplistic liftOver
  call       call variants from asm-to-ref alignment with the cs tag
  bedcov     compute the number of bases covered

  mapeval    evaluate mapping accuracy using mason2/PBSIM-simulated FASTQ
  mason2fq   convert mason2-simulated SAM to FASTQ
  pbsim2fq   convert PBSIM-simulated MAF to FASTQ
  junceval   evaluate splice junction consistency with known annotations
  ov-eval    evaluate read overlap sensitivity using read-to-ref mapping

# 安装
conda create -n hisat2 -c conda-forge -c bioconda hisat2
conda activate hisat2

# 建索引
hisat2-build ref.fa ref.idx -p 20

# 双端测序的比对
hisat2 -q -x ref.idx -1 B2_1.clean.fq.gz -2 B2_2.clean.fq.gz -S out.sam -p 25

--end-to-end 调用bowtie2的参数全长比对

STAR --runThreadN 20 # 线程数 
--runMode genomeGenerate 
--genomeDir star_index/ # index输出的路径 （事先准备好的index文件夹）
--genomeFastaFiles ref.fa  # 参考基因组（之前下载的.fa文件）
--sjdbGTFfile gencode.v38.annotation.gtf  # 参考基因组注释文件 （之前下载的.gtf文件）
--sjdbOverhang 35 # readlength-1 (默认值是100）,read长度-1

将bam文件和对应的bai文件拷到机器

项目地址：https://github.com/lh3/bwa 及 https://github.com/bwa-mem2/bwa-mem2。能够将差异度较小的短序列比对到一个较大的参考基因组上

# 安装
conda create -n bwa -c conda-forge -c bioconda bwa
conda activate bwa
conda create -n bwa-mem2 -c conda-forge -c bioconda bwa-mem2
conda activate bwa-mem2

# 建索引
bwa-mem2 index ref.fa

# 比对
bwa-mem2 mem -t <num_threads> ref.fa read1.fq read2.fq > out.sam

项目地址：https://github.com/samtools/samtools

# 安装
conda create -n samtools -c conda-forge -c bioconda samtools
conda activate samtools

比如用于筛选叶绿体基因组三代reads

项目地址：https://github.com/CMU-SAFARI/BLEND。类似minimap2

2023-11-02

reads比对

minimap2

paftools.js

hisat2：rna-seq比对，快速、低内存

STAR：RNA-Seq reads

IGV：reads比对可视化

jBrowse2

bwa & bwa-mem2：二代比对到参考基因组

samtools

bstr

BLEND

本页目录