{"id":1604,"date":"2013-06-16T15:20:10","date_gmt":"2013-06-16T07:20:10","guid":{"rendered":"http:\/\/www.hzaumycology.com\/chenlianfu_blog\/?p=1604"},"modified":"2013-06-16T21:18:27","modified_gmt":"2013-06-16T13:18:27","slug":"%e8%ae%a1%e7%ae%97paired-end%e6%b5%8b%e5%ba%8f%e7%9a%84insert-size","status":"publish","type":"post","link":"http:\/\/www.chenlianfu.com\/?p=1604","title":{"rendered":"\u8ba1\u7b97paired-end\u6d4b\u5e8f\u7684insert size"},"content":{"rendered":"<p>\u7ed9\u51fa\u7684\u6761\u4ef6\u4e3a\uff1a<br \/>\n\u53c2\u8003\u5e8f\u5217\u6587\u4ef6\uff1a ref.fasta<br \/>\nIllumnina paired-end\u6587\u4ef6\uff1a reads1.fq, reads2.fq<\/p>\n<h1>1. \u7ed3\u5408 Bowtie2\uff0cSAMtools \u548c Picard\u7684\u4f7f\u7528\u6765\u8ba1\u7b97insert size<\/h1>\n<h2>1.1 \u5c06paired-end\u6570\u636e\u6bd4\u5bf9\u5230\u53c2\u8003\u5e8f\u5217\u4e0a<\/h2>\n<pre>\r\n$ bowtie2-build ref.fasta ref\r\n$ bowtie2 --rg-id librarylength_insert --rg \"PL:ILLUMINA\" \\\r\n  --rg \"SM:Strain\" -x ref -p 8 \\\r\n  -1 reads1.fq -2 reads2.fq -S align.sam\r\n<\/pre>\n<h2>1.2 \u5c06sam\u683c\u5f0f\u6bd4\u5bf9\u7ed3\u679c\u8f6c\u6362\u4e3abam\u683c\u5f0f\u5e76\u6392\u5e8f<\/h2>\n<p>\u4f7f\u7528samtools\u5c06sam\u6587\u4ef6\u8f6c\u6362\u6210bam\u6587\u4ef6\uff1b\u4f7f\u7528picard\u5c06bam\u6587\u4ef6\u8fdb\u884c\u6392\u5e8f.<\/p>\n<pre>\r\n$ samtools view -bS align.sam > align.bam\r\n$ jar Xmx2g -jar $picardHome\/SortSam.jar \\\r\n  I=align.bam O=align.sort.bam SO=coordinate\r\n<\/pre>\n<h2>1.3 \u8ba1\u7b97insertsize<\/h2>\n<p>\u8f93\u5165\u6392\u5e8f\u540e\u7684bam\u6587\u4ef6\uff0c\u4f7f\u7528picard\u751f\u6210insertsize\u7684txt\u7684\u7ed3\u679c\u6587\u4ef6\u548cPDF\u683c\u5f0f\u7684\u56fe\u5f62\u7ed3\u679c\u3002<\/p>\n<pre>\r\n$ jar Xmx2g -jar $picardHome\/CollectInsertSizeMetrics.jar \\\r\n  I=align.sort.bam R=ref.fasta  \\\r\n  O=insertsize.txt H=insertsize.pdf\r\n<\/pre>\n<h1> 2. \u4f7f\u7528Qualimap\u6765\u8ba1\u7b97insertsize<\/h1>\n<h2>2.1 Qualimap\u7684\u4ecb\u7ecd<\/h2>\n<p><a href=\"http:\/\/qualimap.bioinfo.cipf.es\/\" target=\"_blank\">Qualimap<\/a> \u7528\u4e8eNGS\u6bd4\u5bf9\u6570\u636e\u7684\u8d28\u91cf\u63a7\u5236\u3002\u4f7f\u7528JAVA\u548cR\u7f16\u5199\u7684\u7a0b\u5e8f\uff0c\u6709GUI\u548ccommand-line\u4e24\u79cd\u8fd0\u884c\u65b9\u5f0f\u3002<\/p>\n<p>\u6587\u732e\uff1aFernando Garc\u00eda-Alcalde, Konstantin Okonechnikov, Jos\u00e9 Carbonell, Luis M. Cruz, Stefan G\u00f6tz, Sonia Tarazona, Joaqu\u00edn Dopazo, Thomas F. Meyer, and Ana Conesa &#8220;Qualimap: evaluating next-generation sequencing alignment data.&#8221; Bioinformatics 28, no. 20 (2012): 2678-2679.<br \/>\n<a href=\"http:\/\/bioinformatics.oxfordjournals.org\/content\/28\/20\/2678\" target=\"_blank\">doi: 10.1093\/bioinformatics\/bts503<\/a><\/p>\n<h2>2.2 Qualimap\u7684\u5b89\u88c5<\/h2>\n<p>Qualimap\u7684\u6b63\u5e38\u4f7f\u7528\u9700\u8981 JAVA runtime vesion 6 \u6216\u4ee5\u4e0a\u7248\u672c\uff1b R 2.14 \u6216\u4ee5\u4e0a\u7248\u672c\uff1b\u9700\u8981\u4e00\u4e9b R \u5305\u3002\u8be5\u8f6f\u4ef6\u9ed8\u8ba4\u4f7f\u7528\u6700\u9ad8\u7ebf\u7a0b\u6570\u8fd0\u884c\uff0c\u901f\u5ea6\u4e0d\u9519\u3002<\/p>\n<pre>\r\n$ wget http:\/\/qualimap.bioinfo.cipf.es\/release\/qualimap_v0.7.1.zip\r\n$ unzip qualimap_v0.7.1.zip\r\n$ cd qualimap_v0.7.1\r\n$ sudo su\r\n$ \/usr\/local\/bin\/Rscript scripts\/installDependencies.r\r\n$ R\r\n&gt; source(\"http:\/\/bioconductor.org\/biocLite.R\")\r\n&gt; biocLite(\"gplots\",\"Repitools\")\r\n<\/pre>\n<h1>3. \u6ce8\u610f\u4e8b\u9879<\/h1>\n<p>\u5bf9\u4e8e\u57fa\u56e0\u7ec4\u7684paired-end\u6d4b\u5e8f\uff0c\u5c06reads\u6bd4\u5bf9\u5230\u53c2\u8003\u57fa\u56e0\u7ec4\u4e0a\uff1b\u4f46\u662f\u5bf9\u4e8e\u8f6c\u5f55\u7ec4\u6d4b\u5e8f\u7684paired-end\u6d4b\u5e8f\uff0c\u5219\u6700\u597d\u5c06reads\u6bd4\u5bf9\u5230\u8f6c\u5f55\u7ec4\u5e8f\u5217\u4e0a\u3002\u800c\u6bd4\u5bf9\u5230\u57fa\u56e0\u7ec4\u5e8f\u5217\u4e0areads\u6570\u4f1a\u504f\u5c11\u3002<\/p>\n","protected":false},"excerpt":{"rendered":"<p>\u7ed9\u51fa\u7684\u6761\u4ef6\u4e3a\uff1a \u53c2\u8003\u5e8f\u5217\u6587\u4ef6\uff1a ref.fasta Illumnina pair &hellip; <a href=\"http:\/\/www.chenlianfu.com\/?p=1604\">\u7ee7\u7eed\u9605\u8bfb <span class=\"meta-nav\">&rarr;<\/span><\/a><\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":[],"categories":[],"tags":[],"_links":{"self":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/1604"}],"collection":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=1604"}],"version-history":[{"count":8,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/1604\/revisions"}],"predecessor-version":[{"id":1621,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/1604\/revisions\/1621"}],"wp:attachment":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=1604"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=1604"},{"taxonomy":"post_tag","embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=1604"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}