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共线性分析——MCScanX

2019-01-09  本文已影响115人  bioinfo_boy

目录

  • MCScanX下载及安装
  • 数据准备
  • MCScanX运行及结果
  • Ka & Ks 计算
  • 下游画图脚本
  • 最后

MCScanX有两个主要的功能, 一是, 方便使用者发现共线性(collinearity)和同线性(synteny)关系并且可以从共线性区块中看到清晰多重比对; 二是, 通过其中众多的辅助脚本, 更形象的分析同线性和共线性的数据
下面就从零开始共线性的分析吧~
注: 稍微解释一下synteny and collinearity的关系, 假设有A B C三个基因, 在species1中的排列为ACB, 而在species2中排列为CAB, 则这两个区段称为synteny; 如果在species3中排列为ABC, 在species4中排列也为ABC, 则这两个区段称为collinearity

MCScanX下载及安装

$wget http://chibba.pgml.uga.edu/mcscan2/MCScanX.zip
$unzip MCScanX.zip
$cd MCScanX
$make

这部分我是在华为云服务器上完成的, 因为很多东西没装, 所以一直报错, 报错内容及解决办法记录如下:

make: g++: Command not found

$conda install gcc=4.8.5

g++ struct.cc mcscan.cc read_data.cc out_utils.cc dagchainer.cc msa.cc permutation.cc -o MCScanX
msa.cc: In function ‘void msa_main(const char*)’:
msa.cc:289:22: error: ‘chdir’ was not declared in this scope
     if (chdir(html_fn)<0)
                      ^
makefile:2: recipe for target 'mcscanx' failed
make: *** [mcscanx] Error 1

#include <unistd.h>

can't find javac

$yum -y install  java-1.8.0-openjdk-devel.x86_64

数据准备

安装完成后, 软件包包括三个主程序MCScanX, MCScanX_h, duplicate_gene_classifier, 和13个用于形象化展示的下游分析软件
本次分析只用到了MCScanX和几个下游脚本

BLASTP

$makeblastdb -in pde_ptr_pep.fasta -dbtype prot -parse_seqids -out pep.db/pde_ptr

#PBS -N pdeptr_blastp
#PBS -l nodes=1:ppn=62
#PBS -l walltime=1200:00:00
#PBS -q long
cd /home/Dai_XG/Analysis/Pdel_genome/05.evolution_analysis/05.colinearity/out_01
blastp -query \pde_ptr_pep.fasta -out pde_ptr_pep.fasta.blastpout -db pep.db/pdeptr -outfmt 6 -evalue 1e-5 -num_threads 62

特定GFF生成

需要的格式只有四列信息, 分别是染色体ID, 基因ID, 起始和终止, 以下命令行是对基因注释文件操作的
如果只想做特定序列的结果, 在这一步中筛出来就行, 不用动上边的 blast 结果

#具体文件具体分析
$grep '\smRNA\s' EVM.final.gene.gff3 | awk '{print $1"\t"$4"\t"$5"\t"$9}' |awk -F 'ID=' '{print $1$2}'|awk -F 'Parent=' '{print $1}'|awk '{print $1"\t"$4"\t"$2"\t"$3}' >pde.gff 
$grep '\smRNA\s' Ptrichocarpa_444_v3.1.gene.gff3 | awk '{print $1"\t"$4"\t"$5"\t"$9}'|awk -F '.v3.1;' '{print $1}'|awk -F 'ID=' '{print $1$2}'|awk '{print $1"\t"$4"\t"$2"\t"$3}' > ptr.gff

MCScanX运行及结果

运行命令行

$nohup MCScanX pde_ptr > out.log 2>&1 &

结果

运行完成后会生成三个结果文件

pde_ptr.collinearity

就不粘自己电脑上的图了, 偷张别人的图说一下

pde_ptr.html

目录中包含所有 block 的.html 文件

pde_ptr.tandem

串联重复的基因, 也不知道有啥用

Ka & Ks 计算

上边说了, MCScanX有很多下游分析脚本, 我这几篇文章都是在记录之前做的进化分析流程, 其中流程中做过KaKs的统计计算, 下面就着重介绍一下, 关于其他下游脚本, 将在下一章节介绍

定义

数据准备

软件配置

这一步需要下游脚本add_ka_and_ks_to_synteny.pl, 如果你也是普通用户, 那么就很操蛋了, 要手动下载安装很多个perl model, 还要装ClustalX, 而且必须是2.0版本以前的

$wget http://www.clustal.org/download/1.X/ftp-igbmc.u-strasbg.fr/pub/ClustalW/clustalw1.83.linux.tar.gz
$tar zxvf clustalw1.83.linux.tar.gz
$vi ~/.bash_profile

export PATH=/home/Dai_XG/software/clustalw1.83.linux:$PATH

$wget https://cpan.metacpan.org/authors/id/C/CD/CDRAUG/Bio-Tools-Run-Alignment-Clustalw-1.7.4.tar.gz
$tar zxvf Bio-Tools-Run-Alignment-Clustalw-1.7.4.tar.gz
$cd Bio-Tools-Run-Alignment-Clustalw
$perl Makefile.PL PREFIX=/home/Dai_XG/software/prefix/perl_local_lib
$make
$make test
$make install

$wget https://cpan.metacpan.org/authors/id/T/TO/TOKUHIROM/Devel-CheckBin-0.04.tar.gz
$tar zxvf Devel-CheckBin-0.04.tar.gz
$cd Devel-CheckBin-0.04
$perl Makefile.PL PREFIX=/home/Dai_XG/software/prefix/perl_local_lib
$make
$make test
$make install

$wget https://cpan.metacpan.org/authors/id/C/CJ/CJFIELDS/BioPerl-Run-1.007002.tar.gz
$tar zxvf BioPerl-Run-1.007002.tar.gz
$cd BioPerl-Run-1.007002

这是Bioperl, 手动形式跟上边的不同, 我是在没找到普通用户的管理方法, 但发现目录下有一个lib 目录, 把它写入环境竟然也能用, 太好了~~

$vi ~/.bash_profile

export PERL5LIB=/home/Dai_XG/software/prefix/perl_local_lib/lib/site_perl/5.28.0/:/home/Dai_XG/software/BioPerl-Run-1.007002/lib/:$PERL5LIB
export PATH=/home/Dai_XG/software/clustalw1.83.linux:$PATH

运行及结果

终于能用了, 太烦了

自编脚本处理

两个脚本太长了, 备份里找吧, 名称为extract_peak.kaks.pl, statistics_block.pl

Chr&LachesisXXgroup
-19990  2318
0       1052
220     819
200     805
...
Chr&Chr
-19990  810
2320    73
2280    73
...
LachesisXXgroup&LachesisXXgroup
-19990  2430
2290    76
2160    67

## BLOCK 1      Chr01   2133    LachesisXXgroup0        1624    Ks
0       Chr01   Potri.001G064150.1      5052138 5053352 -2
1       Chr01   Potri.001G064301.1      5072110 5072703 -2
2       Chr01   Potri.001G065600.1      5191942 5199150 -2
...
0       LachesisXXgroup0        EVM0035433.1    4924636 4927232 -2
1       LachesisXXgroup0        EVM0024093.1    5012566 5014702 -2
2       LachesisXXgroup0        EVM0031968.1    5107185 5112594 -2
...
## BLOCK 2      Chr01   2118    LachesisXXgroup0        1564    Ks
0       Chr01   Potri.001G290600.3      29621454        29622702        -2
1       Chr01   Potri.001G290800.3      29637348        29639177        -2
2       Chr01   Potri.001G291866.1      29721339        29722599        -2
...

BLOCK   species blocklength     colinearity genes       block genes with no colinearity
BLOCK1  Chr01   20138601        1344    789     LachesisXXgroup0        20266103        1344    280
BLOCK2  Chr01   20866914        1135    983     LachesisXXgroup0        21403547        1135    429
BLOCK3  Chr01   5024695 434     330     LachesisXXgroup0        5147359 434     157
BLOCK4  Chr01   3688015 309     201     LachesisXXgroup0        4407659 309     86
BLOCK5  Chr01   1523936 21      91      LachesisXXgroup0        1523936 21      62
BLOCK6  Chr01   226943  10      25      LachesisXXgroup0        404525  10      36
BLOCK7  Chr01   308614  9       25      LachesisXXgroup0        308614  9       8
BLOCK8  Chr01   929669  10      71      LachesisXXgroup0        1682958 10      26
BLOCK9  Chr01   1606860 10      104     LachesisXXgroup0        2277577 10      123
BLOCK10 Chr01   38729   6       2       LachesisXXgroup0        143832  6       0
BLOCK11 Chr01   385605  7       44      LachesisXXgroup0        6302890 7       55
BLOCK12 Chr01   80612   6       6       LachesisXXgroup0        42799884        6       30
...

下游画图脚本

这里尽可能多的吧脚本都跑一遍吧, 实在没法跑的以后补充
每个脚本运行前需要.java 及.class 同时存在才可以

bar_plotter.java

2000    //dimension (in pixels) of x axis
2000    //dimension (in pixels) of y axis
Chr01,Chr02,Chr03,Chr04,Chr05,Chr06,Chr07,Chr08,Chr09,Chr10,Chr11,Chr12,Chr13,Chr14,Chr15,Chr16,Chr17,Chr18,Chr19       //reference chromosomes
Lachesis_group0,Lachesis_group1,Lachesis_group2,Lachesis_group3,Lachesis_group4,Lachesis_group5,Lachesis_group6,Lachesis_group7,Lachesis_group8,Lachesis_group9,Lachesis_group10,Lachesis_group11,Lachesis_group12,Lachesis_group13,Lachesis_group14,Lachesis_group15,Lachesis_group16,Lachesis_group17,Lachesis_group18        //target chromosomes

$java dot_plotter -g gff_file -s collinearity_file -c control_file -o output_PNG_file

circle_plotter.java

800     //plot width and height (in pixels)
Chr01,Chr02,Chr03,Chr04,Chr05,Chr06,Chr07,Chr08,Chr09,Chr10,Chr11,Chr12,Chr13,Chr14,Chr15,Chr16,Chr17,Chr18,Chr19,Lachesis_group0,Lachesis_group1,Lachesis_group2,Lachesis_group3,Lachesis_group4,Lachesis_group5,Lachesis_group6,Lachesis_group7,Lachesis_group8,Lachesis_group9,Lachesis_group10,Lachesis_group11,Lachesis_group12,Lachesis_group13,Lachesis_group14,Lachesis_group15,Lachesis_group16,Lachesis_group17,Lachesis_group18  //chromosomes in the circle

$java circle_plotter -g gff_file -s collinearity_file -c control_file -o output_PNG_file

dot_plotter.java

800     //dimension (in pixels) of x axis
800     //dimension (in pixels) of y axis
sb1,sb2,sb3,sb4,sb5,sb6,sb7,sb8,sb9,sb10        //chromosomes in x axis
os1,os2,os3,os4,os5,os6,os7,os8,os9,os10,os11,os12      //chromosomes in y axis

$java dot_plotter -g gff_file -s collinearity_file -c control_file -o output_PNG_file

dual_synteny_plotter.java

6000    //plot width (in pixels)
20000   //plot height (in pixels)
Chr01,Chr02,Chr03,Chr04,Chr05,Chr06,Chr07,Chr08,Chr09,Chr10,Chr11,Chr12,Chr13,Chr14,Chr15,Chr16,Chr17,Chr18,Chr19,      //chromosomes in the left column
Lachesis_group0,Lachesis_group1,Lachesis_group2,Lachesis_group3,Lachesis_group4,Lachesis_group5,Lachesis_group6,Lachesis_group7,Lachesis_group8,Lachesis_group9,Lachesis_group10,Lachesis_group11,Lachesis_group12,Lachesis_group13,Lachesis_group14,Lachesis_group15,Lachesis_group16,Lachesis_group17,Lachesis_group18,       //chromosomes in the right column

$java dual_synteny_plotter -g gff_file -s collinearity_file -c control_file -o output_PNG_file

最后

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