标签：生物信息学

各类生物软件应用心得

magrittr – Ceci n’est pas un pipe.

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tophat library type 应该怎么设置

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当我们拿到一个测序结果，实验人员也搞不清楚他的结果的library type怎么运行tophat呢？

依照手册，你可以先访问实验人员，他使用的是什么样的测序手段，是用的illumina还是SOLiD。如果是SOLiD，多半是second strand. 如果是illumina，可能还是分辨一下，主要是分辨unstraned还是firststrand.

这可能需要分三步走：

1 r[……]

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使用Biostrings的PDict搜索一系列小片段

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今天遇到一个小问题，在给定的一些序列中，搜索CRISPR Toolbox中GeCKO library中已有的guideRNA。这正好符合PDict的要求，PDict要求所有的片段长度必须一致，而guide sequence的长度正好都是20。

代码如下：

> library(Biostrings)
> seq <- dir("tobeSearched", pattern=".seq$")
> seq
 [1] "1-F_A01_015_2015-03-06.seq"  "1-R_C02_012_2015-03-06.seq"  "10-F_B02_014_2015-03-06.seq"
 [4] "10-R_D03_025_2015-03-06.seq" "2-F_B01_013_2015-03-06.seq"  "2-R_D02_010_2015-03-06.seq" 
 [7] "3-F_C01_011_2015-03-06.seq"  "3-R_E02_008_2015-03-06.seq"  "4-F_D01_009_2015-03-06.seq" 
[10] "4-R_F02_006_2015-03-06.seq"  "5-F_E01_007_2015-03-06.seq"  "5-R_G02_004_2015-03-06.seq" 
[13] "6-F_F01_005_2015-03-06.seq"  "6-R_H02_002_2015-03-06.seq"  "7-F_G01_003_2015-03-06.seq" 
[16] "7-R_A03_031_2015-03-06.seq"  "8-F_H01_001_2015-03-06.seq"  "8-R_B03_029_2015-03-06.seq" 
[19] "9-F_A02_016_2015-03-06.seq"  "9-R_C03_027_2015-03-06.seq" 
> sequences <- sapply(seq, function(.ele){ readLines(file.path("tobeSearched", .ele))})
> sequences[1:2]
$`1-F_A01_015_2015-03-06.seq`
 [1] "NNNNNNNNNNNNNNNNNNNNNNNNTNNNNNNNGNNNNNNNNNNNNNNNNNNNNNNNAATATCCCCCNGACGGNNNGNGNN"
 [2] "NNNNNNNNNNNNTTCAGGCTGNNNNNNNCACNNNNNNCGAGNCATCANNNNNAGGGNGNGNTTGNNNCTCANNNNNNCAG"
 [3] "GTNCTGNNNGNNGCGTTNAGTGNCATCNCCGGTNGGNNNNNNNNNGGNNAAACCNCNCGNANNCCCNNCNNGAAGCNNNN"
 [4] "ACNTTNNTTNNNCCCNANNNGCCCNAAATGTTTNANCAANANNTNNNNNNNNNNNGNNNNCTTNTNNTNNNNNNNNNNNN"
 [5] "NNNNNNGNNNNNCGGNNGGNACNAANCNNCCNNNNNNNNANCANANGCNGGGGNNAAATTTNNCCCCNNCANNNNNNNNA"
 [6] "ANGGGNNCCCCNGNAANNTTTTTNNNAANNNCTTTCCCCCCCCNNAGGCCTTTCCCCCNNNAANNCCNCCCCCNNNNNNN"
 [7] "NNNNNGGNAAAAAAAAAAAAAAANNNNNNNAAAAANNNNGNNNNNNNNNNNAAAAAGNATNNNTNNTGNNNNNNNNNNTG"
 [8] "NNNNNNNNNANNNNAANNNNNNNNNTTNNGAANNANNGNNNGNNNNNNNNGTNNTANNNNNGNGNNTATTNNGNTTANNG"
 [9] "GNNNNGNGNNTNNNNNNNNGNGNNNNNNNNCNGTNNNTNNNNNNNTNNNNNNNNNNGNNGNNGTNGNNGTNNNNNNNGNN"
[10] "NNGGNNNNNNGNNGGGNGNNNGGNNNNNNNNNGNCNNNTNNNNNNGNGTNGNNGNGNNNGNNNNNTANTNTNTNNTAANA"
[11] "ANGNGNNNNGTGNTNTTTNNNNNNNTNNNNNNNNNNNNNNNNNNNNNNGNNNTTNNNNNNNNNNNNNNNNNN"        
 
$`1-R_C02_012_2015-03-06.seq`
[1] "NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNCCGGCNNAAGNCATTATGCTCGGNNAAAGNACTTCCTGGCGCGTGNTTAC"
[2] "CAGGGCGCACCGATTGCCATCACCGTTGAAGGGGCTAACATTCTGACCCGCNGCATGATGNTCTTCGGACAAGGNGCGAT"
[3] "TCGTTGCCATCCGTACGTGCTGGANGAGANGGAAGCGGCGAANAANAATGANGTCAACGCGTTCGATAAACTGTTGTTCA"
[4] "AACATATCGGTCACGNCGGTAGCAACAAAGNTCGCAGCTTCTGGCTGGNCNNGANGCGCGGNTTAACCAGCAGCACGNCA"
[5] "ACCGGCGATGCCACTAAACGCTACTATCAGCACCTGANCNGCNGAGCGCCNNCCNCGCNNTGNTNNTGANGNNTCGNNNN"
[6] "NGNNNNGGNGNNNNNNAANNNCNNNNNNNNNNGNNNNNTTNTTTNNNNNNANNCNNCCCNCCNNNNNNNCNGNNNNNNNN"
[7] "NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNGNNNNNNN"                                
 
> sequences <- lapply(sequences, paste, collapse="")
> sequences <- unlist(sequences)
> sequences[1]
                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                1-F_A01_015_2015-03-06.seq 
"NNNNNNNNNNNNNNNNNNNNNNNNTNNNNNNNGNNNNNNNNNNNNNNNNNNNNNNNAATATCCCCCNGACGGNNNGNGNNNNNNNNNNNNNNTTCAGGCTGNNNNNNNCACNNNNNNCGAGNCATCANNNNNAGGGNGNGNTTGNNNCTCANNNNNNCAGGTNCTGNNNGNNGCGTTNAGTGNCATCNCCGGTNGGNNNNNNNNNGGNNAAACCNCNCGNANNCCCNNCNNGAAGCNNNNACNTTNNTTNNNCCCNANNNGCCCNAAATGTTTNANCAANANNTNNNNNNNNNNNGNNNNCTTNTNNTNNNNNNNNNNNNNNNNNNGNNNNNCGGNNGGNACNAANCNNCCNNNNNNNNANCANANGCNGGGGNNAAATTTNNCCCCNNCANNNNNNNNAANGGGNNCCCCNGNAANNTTTTTNNNAANNNCTTTCCCCCCCCNNAGGCCTTTCCCCCNNNAANNCCNCCCCCNNNNNNNNNNNNGGNAAAAAAAAAAAAAAANNNNNNNAAAAANNNNGNNNNNNNNNNNAAAAAGNATNNNTNNTGNNNNNNNNNNTGNNNNNNNNNANNNNAANNNNNNNNNTTNNGAANNANNGNNNGNNNNNNNNGTNNTANNNNNGNGNNTATTNNGNTTANNGGNNNNGNGNNTNNNNNNNNGNGNNNNNNNNCNGTNNNTNNNNNNNTNNNNNNNNNNGNNGNNGTNGNNGTNNNNNNNGNNNNGGNNNNNNGNNGGGNGNNNGGNNNNNNNNNGNCNNNTNNNNNNGNGTNGNNGNGNNNGNNNNNTANTNTNTNNTAANAANGNGNNNNGTGNTNTTTNNNNNNNTNNNNNNNNNNNNNNNNNNNNNNGNNNTTNNNNNNNNNNNNNNNNNN" 
 
> sequences <- DNAStringSet(sequences)
> head(sequences)
  A DNAStringSet instance of length 6
    width seq                                                                  names               
[1]   872 NNNNNNNNNNNNNNNNNNNNNNNNTNNNNNNNG...NNNNNNNNGNNNTTNNNNNNNNNNNNNNNNNN 1-F_A01_015_2015-...
[2]   528 NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNCCG...NNNNNNNNNNNNNNNNNNNNNNNNGNNNNNNN 1-R_C02_012_2015-...
[3]   543 NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNG...NNNNNNNNNNNNTNNNNNNNNNNNNNNNNNNN 10-F_B02_014_2015...
[4]   525 GNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNC...NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN 10-R_D03_025_2015...
[5]   697 NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN...NNNGNNNNNNNNNCTTTTTTTTTTNNNNNNAN 2-F_B01_013_2015-...
[6]   527 NNNNNNNNNNNNNNNNNNNNNNNNGNNNNNNNC...NNNNNNNNNNNNNNNNNNNNNNNNNNAANNNN 2-R_D02_010_2015-...
> library1 <- read.csv("mouse_geckov2_library_b_1.10000000053.csv", stringsAsFactors=F)
> library2 <- read.csv("mouse_geckov2_library_a_2.10000000053.csv", stringsAsFactors=F)
> head(library1)
        gene_id          UID                  seq
1 0610007P14Rik MGLibB_00001 TGTCTAAGGTTTCTCAATCC
2 0610007P14Rik MGLibB_00002 CAGTGAATGGCCTCCAAGCC
3 0610007P14Rik MGLibB_00003 GGTGCTTACTTGCTACCATC
4 0610009B22Rik MGLibB_00004 ACCTCGTCGACGAAAACATG
5 0610009B22Rik MGLibB_00005 GTTCATAGCTCATGCTGCTC
6 0610009B22Rik MGLibB_00006 GAGCAGCATGAGCTATGAAC
> head(library2)
        gene_id          UID                  seq
1 0610007P14Rik MGLibA_00001 TCCTGAATGTGTTACGAAGC
2 0610007P14Rik MGLibA_00002 GGTCGGGCTCCGGTACCTAG
3 0610007P14Rik MGLibA_00003 GCCAGCTTCGTAACACATTC
4 0610009B22Rik MGLibA_00004 TCATCATGCTGCATGACGTG
5 0610009B22Rik MGLibA_00005 ATTCAATGAGTGGTTCGTCT
6 0610009B22Rik MGLibA_00006 TCGTCACGGCTGGGCACATG
> dim(library1)
[1] 62804     3
> dim(library2)
[1] 67405     3
> library1.DNA <- library1$seq
> library2.DNA <- library2$seq
> names(library1.DNA) <- library1$UID
> names(library2.DNA) <- library2$UID
> library1.DNA <- DNAStringSet(library1.DNA)
> library2.DNA <- DNAStringSet(library2.DNA)
> head(library1.DNA)
  A DNAStringSet instance of length 6
    width seq                                                                  names               
[1]    20 TGTCTAAGGTTTCTCAATCC                                                 MGLibB_00001
[2]    20 CAGTGAATGGCCTCCAAGCC                                                 MGLibB_00002
[3]    20 GGTGCTTACTTGCTACCATC                                                 MGLibB_00003
[4]    20 ACCTCGTCGACGAAAACATG                                                 MGLibB_00004
[5]    20 GTTCATAGCTCATGCTGCTC                                                 MGLibB_00005
[6]    20 GAGCAGCATGAGCTATGAAC                                                 MGLibB_00006
> head(library2.DNA)
  A DNAStringSet instance of length 6
    width seq                                                                  names               
[1]    20 TCCTGAATGTGTTACGAAGC                                                 MGLibA_00001
[2]    20 GGTCGGGCTCCGGTACCTAG                                                 MGLibA_00002
[3]    20 GCCAGCTTCGTAACACATTC                                                 MGLibA_00003
[4]    20 TCATCATGCTGCATGACGTG                                                 MGLibA_00004
[5]    20 ATTCAATGAGTGGTTCGTCT                                                 MGLibA_00005
[6]    20 TCGTCACGGCTGGGCACATG                                                 MGLibA_00006
> library1.PDict <- PDict(library1.DNA)
> library2.PDict <- PDict(library2.DNA)
> mT.1 <- lapply(sequences, function(.ele){ whichPDict(library1.PDict, .ele)})
> mT.2 <- lapply(sequences, function(.ele){ whichPDict(library2.PDict, .ele)})
> sapply(mT.1, length)
 1-F_A01_015_2015-03-06.seq  1-R_C02_012_2015-03-06.seq 10-F_B02_014_2015-03-06.seq 
                          0                           0                           0 
10-R_D03_025_2015-03-06.seq  2-F_B01_013_2015-03-06.seq  2-R_D02_010_2015-03-06.seq 
                          0                           0                           0 
 3-F_C01_011_2015-03-06.seq  3-R_E02_008_2015-03-06.seq  4-F_D01_009_2015-03-06.seq 
                          0                           0                           0 
 4-R_F02_006_2015-03-06.seq  5-F_E01_007_2015-03-06.seq  5-R_G02_004_2015-03-06.seq 
                          0                           0                           0 
 6-F_F01_005_2015-03-06.seq  6-R_H02_002_2015-03-06.seq  7-F_G01_003_2015-03-06.seq 
                          0                           0                           0 
 7-R_A03_031_2015-03-06.seq  8-F_H01_001_2015-03-06.seq  8-R_B03_029_2015-03-06.seq 
                          1                           0                           0 
 9-F_A02_016_2015-03-06.seq  9-R_C03_027_2015-03-06.seq 
                          0                           0 
> sapply(mT.2, length)
 1-F_A01_015_2015-03-06.seq  1-R_C02_012_2015-03-06.seq 10-F_B02_014_2015-03-06.seq 
                          0                           0                           0 
10-R_D03_025_2015-03-06.seq  2-F_B01_013_2015-03-06.seq  2-R_D02_010_2015-03-06.seq 
                          0                           0                           0 
 3-F_C01_011_2015-03-06.seq  3-R_E02_008_2015-03-06.seq  4-F_D01_009_2015-03-06.seq 
                          0                           0                           0 
 4-R_F02_006_2015-03-06.seq  5-F_E01_007_2015-03-06.seq  5-R_G02_004_2015-03-06.seq 
                          0                           0                           0 
 6-F_F01_005_2015-03-06.seq  6-R_H02_002_2015-03-06.seq  7-F_G01_003_2015-03-06.seq 
                          0                           1                           0 
 7-R_A03_031_2015-03-06.seq  8-F_H01_001_2015-03-06.seq  8-R_B03_029_2015-03-06.seq 
                          0                           0                           0 
 9-F_A02_016_2015-03-06.seq  9-R_C03_027_2015-03-06.seq 
                          0                           0

如何使用biomaRt做hg19注释

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因为现在ensembl已经更新了human的注释库，所以当你直接使用

library("biomaRt")
database <- listDatasets(useMart("ensembl"))
database[database[,1]=='hsapiens_gene_ensembl',]
                 dataset                 description version
32 hsapiens_gene_ensembl Homo sapiens genes (GRCh38)  GRCh38

你会得到的是GRCh38（hg38）的注释库。
那么如何转换至hg19注释库呢？
使用

mart <- useMart("ENSEMBL_MART_ENSEMBL", host="grch37.ensembl.org", path="/biomart/martservice")
database <- listDatasets(mart)
database[database[,1]=='hsapiens_gene_ensembl',]

[……]

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R/Bioconductor开发中import和depend的区别

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import:将命名空间载入

depends:将命名空间载入，并且放入搜索目录。

两者在使用上的区别是：import只载入对开发者有用的，而depends需要载入对用户有用的。比如说GenomicRanges，
如果你开发的包使用到了GRanges类，并且它会成为结果返回给用户，那么你就必须使用depends了，如果你只是用了GenomicRanges中的类或者函数，比如说GRang[……]

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