最近接二連三帶了不少實(shí)習(xí)生和輪轉(zhuǎn)生馁启，可以預(yù)見后面幾年實(shí)驗(yàn)室再有實(shí)習(xí)或者輪轉(zhuǎn)的十有八九應(yīng)該都是我?guī)А?br> 這一篇列舉一些生物信息部分常用工具和幾個(gè)神奇網(wǎng)站驾孔。基本上每個(gè)工具都給出一兩句（或中文或英文）簡(jiǎn)要功能介紹和官網(wǎng)地址惯疙。
師妹翠勉，你要的，都在這里了霉颠。

生物信息學(xué)常用工具

fastq格式相關(guān)

• SRAtoolkit

  • SRA數(shù)據(jù)庫(kù)下載公用數(shù)據(jù)時(shí)的工具
  • https://trace.ncbi.nlm.nih.gov/Traces/sra/sra.cgi?view=toolkit_doc

• fastx toolkit

  • a collection of command line tools for Short-Reads FASTA/FASTQ files preprocessing
  • 有各種各樣的小功能对碌，比如提取反向互補(bǔ)序列等等。
  • http://hannonlab.cshl.edu/fastx_toolkit/

• fastqc

  • A quality control tool for high throughput sequence data
  • 評(píng)估測(cè)序數(shù)據(jù)質(zhì)量
  • https://www.bioinformatics.babraham.ac.uk/projects/fastqc/

• MultQC

  • Aggregate results from bioinformatics analyses across many samples into a single report
  • 一次同時(shí)生成多個(gè)數(shù)據(jù)質(zhì)量報(bào)告掉分，省時(shí)省力方便對(duì)比俭缓，支持fastqc
  • https://github.com/ewels/MultiQC; http://multiqc.info/docs/

• Trim Galore

  • around Cutadapt and FastQC to consistently apply quality and adapter trimming to FastQ files,
  • with some extra functionality for MspI-digested RRBS-type (Reduced Representation Bisufite-Seq) libraries
  • 和fastqc出自一家，可以和fastqc結(jié)合使用酥郭，用來清洗原始數(shù)據(jù)华坦。
  • https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/

• Trimmomatic

  • A flexible read trimming tool for Illumina NGS data
  • 專門清洗illumina測(cè)序數(shù)據(jù)的工具
  • http://www.usadellab.org/cms/index.php?page=trimmomatic

• khmer

  • working with DNA shotgun sequencing data from genomes, transcriptomes, metagenomes, and single cells.
  • 可以對(duì)原始測(cè)序數(shù)據(jù)進(jìn)行過濾等
  • http://khmer.readthedocs.io/en/v2.1.1/user/scripts.htm

BED格式相關(guān)

• bedops
  • 玩轉(zhuǎn)bed格式文件，速度比bedtools快
  • the fast, highly scalable and easily-parallelizable genome analysis toolkit
  • https://bedops.readthedocs.io/en/latest/index.html
• bedtools
  • 最知名的bed文件相關(guān)工具不从，但是和samtools并非出自一家
  • a powerful toolset for genome arithmetic
  • http://bedtools.readthedocs.io/en/latest/index.html

SAM/BAM

• samtools
  • 有這一個(gè)就夠了
  • Utilities for the Sequence Alignment/Map (SAM) format
  • http://www.htslib.org/doc/samtools.html

SNP（VCF/BCF）格式相關(guān)

• GATK
  • 使用率最高的軟件
  • https://software.broadinstitute.org/gatk/documentation/
• bcftools
  • 對(duì)vcf格式的文件進(jìn)行各種操作
  • utilities for variant calling and manipulating VCFs and BCFs
  • http://www.htslib.org/doc/bcftools.html
• vcftools
  • 和bcftools類似
  • https://vcftools.github.io/man_latest.html
• snpEFF
  • Genetic variant annotation and effect prediction toolbox
  • 適合用來進(jìn)行snp注釋
  • 用法 http://snpeff.sourceforge.net/SnpEff_manual.html
  • http://snpeff.sourceforge.net/
  • 也可以注釋ChIP-seq
  • 支持非編碼注釋惜姐，如組蛋白修飾
• samtools mpileup
  • Utilities for the Sequence Alignment/Map (SAM) format
  • http://www.htslib.org/doc/samtools.html

ChIP-seq/motif

peak calling

• MACS

  • Model-based Analysis of ChIP-Seq
  • 主要用于組蛋白修飾產(chǎn)生的narrow peaks(H3K4me3 and H3K9/27ac)
  • transcription factors which are usually associated with sharp and solated peaks
  • http://liulab.dfci.harvard.edu/MACS/README.html

• MACS2

  • MACS的升級(jí)版本，也可以用來找broad peak
  • https://github.com/taoliu/MACS

• SICER

  • 出來懟MACS椿息，主要用來找一些比較寬的peak,類似于H3K9me3 和 H3K36me3歹袁。
  • highly recommended for a practical ChIP-seq experiment design and can be used to account for local biases resulting from read mappability, DNA repeats, local GC content
  • https://www.genomatix.de/online_help/help_regionminer/sicer.html

• 后續(xù)分析可能會(huì)用到的工具

  img

• MAnorm

  • http://bioinfo.sibs.ac.cn/shaolab/MAnorm/MAnorm.htm

large sequences alignment

長(zhǎng)序列比對(duì)常用的幾個(gè)軟件

• MUMer
  • rapid alignment of very large DNA and amino acid sequences
  • http://mummer.sourceforge.net/examples/
  • http://mummer.sourceforge.net/manual/
• GMAP
  • GMAP: A Genomic Mapping and Alignment Program for mRNA and EST Sequences
  • http://research-pub.gene.com/gmap/
• BLAT
  • Blat produces two major classes of alignments:at the DNA level between two sequences that are of 95% or greater identity, but which may include large inserts；at the protein or translated DNA level between sequences that are of 80% or greater identity and may also include large inserts.
  • https://genome.ucsc.edu/goldenpath/help/blatSpec.html

short reads alignment

短序列比寝优，二代測(cè)序數(shù)據(jù)比對(duì)

• BWA
  • Burrows-Wheeler Alignment Tool
  • mapping low-divergent sequences against a large reference genome
  • It consists of three algorithms: BWA-backtrack, BWA-SW and BWA-MEM.
  • http://bio-bwa.sourceforge.net/bwa.shtml
  • https://github.com/lh3/bwa
• GSNAP:
  • Genomic Short-read Nucleotide Alignment Program
  • http://research-pub.gene.com/gmap/
• Bowtie
  • works best when aligning short reads to large genomes
  • not yet report gapped alignments
  • http://bowtie-bio.sourceforge.net/manual.shtml
• Bowtie2
  • 和上一代的區(qū)別在于支持gapped alignments
  • ultrafast and memory-efficient tool for aligning sequencing reads to long reference sequences
  • supports gapped, local, and paired-end alignment modes
  • http://bowtie-bio.sourceforge.net/bowtie2/manual.shtml#reporting
• HISAT2
  • Tophat的繼任者条舔，基于HISAT和Bowtie2
  • HISAT2的速度比STAR快一些
  • http://ccb.jhu.edu/software/hisat2/manual.shtml
• STAR
  • Spliced Transcripts Alignment to a Reference
  • https://github.com/alexdobin/STAR
  • https://github.com/alexdobin/STAR/blob/master/doc/STARmanual.pdf

genome guide assemble

• stringtie
  • highly efficient assembler of RNA-Seq alignments into potential transcripts
  • 對(duì)于可變剪切的發(fā)現(xiàn)相對(duì)準(zhǔn)確
  • https://ccb.jhu.edu/software/stringtie/
• Cufflinks
  • 基本不用了
• IDP
  • Isoform Detection and Prediction tool
  • gmap+hisat2,也就是長(zhǎng)短序列比對(duì)相結(jié)合，效果不錯(cuò)
  • https://www.healthcare.uiowa.edu/labs/au/IDP/IDP_manual.asp

de novo assemble/gene prediction

下面幾個(gè)軟件結(jié)合起來就是一個(gè)從組裝到注釋再到計(jì)算拼接效率的過程

拼接

• trintiy
  • 傾向于預(yù)測(cè)長(zhǎng)的可變剪接
  • 新版本從之前的過度預(yù)測(cè)越來越傾向于有所保留
  • 比較耗資源乏矾，一般1個(gè)CPU最好分配6G-10G
  • 可以有參或者無參轉(zhuǎn)錄組拼接
  • https://github.com/trinityrnaseq/trinityrnaseq/wiki
• oases
  • 通常得到的N50比較高
  • 檢測(cè)低表達(dá)的基因有一定優(yōu)勢(shì)
  • De novo transcriptome assembler for very short reads
  • https://github.com/dzerbino/oases

注釋

• PASA（內(nèi)包括BLAT和GMAP）
  • 得到拼接好的fasta文件后可以用pasa進(jìn)行基因結(jié)構(gòu)預(yù)測(cè)
  • Gene Structure Annotation and Analysis Using PASA
  • http://pasapipeline.github.io/
• Maker
  • 基因預(yù)測(cè)
  • can be used for de novo annotation of newly sequenced genomes, for updating existing annotations to reflect new evidence, or just to combine annotations, evidence, and quality control statistics
  • http://www.yandell-lab.org/software/maker.html

質(zhì)量檢測(cè)

• TransRate
  • 專業(yè)的拼接質(zhì)量評(píng)估軟件孟抗，有三種評(píng)估模式。
  • reference free quality assessment of de novo transcriptome assemblies
  • http://hibberdlab.com/transrate/
• DETONATE
  • DE novo TranscriptOme rNa-seq Assembly with or without the Truth Evaluation
  • https://genomebiology.biomedcentral.com/articles/10.1186/s13059-014-0553-5
• BUSCO
  • 它的評(píng)估模式和上面兩個(gè)不太一樣
  • based on evolutionarily-informed expectations of gene content from near-universal single-copy orthologs

Estimating transcript abundance

可以分為基于比對(duì)和不基于比對(duì)兩種钻心，其中RSEM和eXpress是基于比對(duì)的凄硼，另外兩種是基于比對(duì)的。

• RSEM
  • RNA-Seq by Expectation-Maximization
  • https://deweylab.github.io/RSEM/README.html
• eXpress
  • quantifying the abundances of a set of target sequences from sampled subsequences
  • https://pachterlab.github.io/eXpress/overview.html
• kallisto
  • 快到飛起
  • 豐度估計(jì)中樣本特異性和讀長(zhǎng)偏好性低
  • quantifying abundances of transcripts from RNA-Seq data
  • https://pachterlab.github.io/kallisto/
• salmon
  • 也是很快
  • quantifying the expression of transcripts using RNA-seq data
  • https://combine-lab.github.io/salmon/

Read count

• htseq-count
  • 數(shù)read, 有它就夠了
  • http://htseq.readthedocs.io/en/release_0.9.1/

Difference expression

和之前的步驟對(duì)應(yīng)捷沸，這里也可以分為基于read數(shù)和基于組裝以及不急于比對(duì)三類工具摊沉。

• limma

  • 用于分析芯片數(shù)據(jù)
  • Linear Models for Microarray Data
  • http://bioconductor.org/packages/release/bioc/html/limma.html

• DEseq

  • http://bioconductor.org/packages/release/bioc/html/DESeq.html

• DEseq2

  • 效果在幾個(gè)工具中相對(duì)好
  • http://bioconductor.org/packages/release/bioc/html/DESeq2.html

• DEGseq

  • Identify Differentially Expressed Genes from RNA-seq data
  • http://www.bioconductor.org/packages/2.6/bioc/html/DEGseq.html

• edgeR

  • Empirical Analysis of Digital Gene Expression Data in R
  • http://www.bioconductor.org/packages/release/bioc/html/edgeR.html

• Ballgown

  • 準(zhǔn)確度有時(shí)不是很好
  • facilitate flexible differential expression analysis of RNA-Seq data
  • organize, visualize, and analyze the expression measurements for your transcriptome assembly.
  • https://github.com/alyssafrazee/ballgown

• sleuth

  • 用來配合kallisto使用
  • https://pachterlab.github.io/sleuth/about

Data visualization

數(shù)據(jù)可視化的工具可以分為本地版本和在線版本

• IGV
  • 本地展示分析結(jié)果的不二選擇
  • Integrative Genomics Viewer
  • http://software.broadinstitute.org/software/igv/
• jbrowse
  • 公開展示數(shù)據(jù)或者給合作者分享時(shí)的不二選擇，快且好看痒给。
  • http://jbrowse.org/code/JBrowse-1.10.2/docs/tutorial/
• DEIVA
  • 差異表達(dá)的可視化在線工具
  • Interactive Visual Analysis of differential gene expression test results
  • http://hypercubed.github.io/DEIVA/
• Heatmapper
  • 用來話各種熱圖的在線工具
  • expression-based heat maps
  • pairwise distance maps
  • correlation maps
  • http://www.heatmapper.ca/
• START
  • 基于shinny的一套R(shí)NA-seq數(shù)據(jù)可視化工具
  • visualize RNA-seq data starting with count data
  • https://kcvi.shinyapps.io/START/

幾個(gè)神奇的網(wǎng)站

• biostars
  • https://www.biostars.org
• R book
  • http://r4ds.had.co.nz/
• python guide
  • http://docs.python-guide.org/en/latest/
• bioptyhon
  • http://biopython.org/DIST/docs/tutorial/Tutorial.html
• Rosalind
  • http://rosalind.info/problems/list-view/
• bioinformatics tools
  • https://omictools.com/
  • https://bioinformatics.ca/links_directory/
• data visualistion catalogue
  • http://datavizcatalogue.com/index.html

暫時(shí)就寫這么多说墨，還有一些自己平時(shí)也很少用的就不放進(jìn)來給他人增加負(fù)擔(dān)了骏全，后面再補(bǔ)充。