{"id":2252,"date":"2014-12-10T23:50:15","date_gmt":"2014-12-10T15:50:15","guid":{"rendered":"http:\/\/www.chenlianfu.com\/?p=2252"},"modified":"2014-12-10T23:50:15","modified_gmt":"2014-12-10T15:50:15","slug":"discovar%e7%9a%84%e4%bd%bf%e7%94%a8","status":"publish","type":"post","link":"http:\/\/www.chenlianfu.com\/?p=2252","title":{"rendered":"DISCOVAR\u7684\u4f7f\u7528"},"content":{"rendered":"

1. DISCOVAR\u7b80\u4ecb<\/h1>\n

DISCOVAR \u662f\u6709 ALLPATHS-LG \u8f6f\u4ef6\u5f00\u53d1\u56e2\u961f\u505a\u51fa\u6765\u7684\u8f6f\u4ef6\u3002\u4e3b\u8981\u7528\u4e8e\u5229\u7528 PE 250bp \u6570\u636e\u4e0e\u53c2\u8003\u57fa\u56e0\u7ec4\u7684\u6bd4\u5bf9\u7ed3\u679c\uff0c\u5bf9\u57fa\u56e0\u7ec4\u8fdb\u884c Variants calling \u7684\u540c\u65f6\uff0c\u8fdb\u884c\u57fa\u56e0\u7ec4\u7684\u7ec4\u88c5\u3002\u7279\u522b\u662f\u8fd1\u671f\u516c\u5e03\u7684 DISCOVAR de novo (experimental) \u8fd8\u80fd\u8fdb\u884c\u57fa\u56e0\u7ec4\u7684 De novo \u7ec4\u88c5\u3002<\/p>\n

2. DISCOVAR\u7684\u4e0b\u8f7d\u548c\u5b89\u88c5<\/h1>\n

2.1 DISCOVAR\u7684\u4e0b\u8f7d\u548c\u5b89\u88c5<\/h2>\n

\u6b64\u8f6f\u4ef6\u7684\u5b89\u88c5\u9700\u8981GCC 4.7\u6216\u4ee5\u4e0a\u7248\u672c\u3002<\/p>\n

\r\n$ wget ftp:\/\/ftp.broadinstitute.org\/pub\/crd\/Discovar\/latest_source_code\/LATEST_VERSION.tar.gz\r\n$ tar zxf LATEST_VERSION.tar.gz\r\n$ cd discov*\r\n$ .\/configure --prefix=\/opt\/biosoft\/discovar && make -j 4 && make install\r\n$ cd ..\r\n$ rm -rf discov* LATEST_VERSION.tar.gz\r\n<\/pre>\n
2.2 DISCOVAR Denovo\u7684\u4e0b\u8f7d\u548c\u5b89\u88c5<\/h2>\n
\u6b64\u8f6f\u4ef6\u7684\u5b89\u88c5\u9700\u8981GCC 4.7\u6216\u4ee5\u4e0a\u7248\u672c\uff0cjemalloc 3.6.0\u6216\u4ee5\u4e0a\u7248\u672c\u548csamtools\uff08\u5982\u679c\u4f7f\u7528bam\u6587\u4ef6\uff0c\u5219\u9700\u8981\uff09\u3002<\/p>\n
\r\n$ wget ftp:\/\/ftp.broadinstitute.org\/pub\/crd\/DiscovarExp\/LATEST_VERSION.tar.gz\r\n$ tar zxf LATEST_VERSION.tar.gz\r\n$ cd discov*\r\n$ sudo yum install *malloc*\r\n\u5982\u679c\u6ca1\u6709\u4e0a\u4e00\u6b65\uff0c\u5219\u5728make\u8fc7\u7a0b\u4e2d\u4f1a\u63d0\u793a\u9519\u8bef\u201c\/usr\/bin\/ld: cannot find -ljemalloc\u201d\r\n$ .\/configure --prefix=\/opt\/biosoft\/discovarDenovo && make -j 4 && make install\r\n$ echo 'export MALLOC_PER_THREAD=1' >> ~\/.bashrc\r\n\u4e0a\u4e00\u6b65\u8bbe\u7f6e\u7528\u4e8eallowing per-threads memory management\uff0c\u80fd\u63d0\u9ad8\u8ba1\u7b97\u6027\u80fd\u3002\r\n$ cd ..\r\n$ rm -rf discov* LATEST_VERSION.tar.gz\r\n<\/pre>\n
3. \u8f6f\u4ef6\u4f7f\u7528\u7684\u6ce8\u610f\u4e8b\u9879<\/h1>\n
1. \u5f3a\u70c8\u63a8\u8350\u4f7f\u7528 PCR-free protocol library<\/a> \u6570\u636e\uff1b\u6570\u636e\u91cf\u63a8\u8350\u4e3a ~60x\uff0c\u7565\u5927\u4e8e\u6216\u5c0f\u4e8e\u8be5\u503c\u4e5f\u662f OK \u7684\u3002
\n2. \u5fc5\u987b\u4f7f\u7528 Illumina MiSeq \u6216 HiSeq 2500 \u6d4b\u5e8f\u4eea\u4ea7\u751f\u7684 >=250 bp \u957f\u5ea6\u7684 Paired End \u6570\u636e\uff0c\u5e76\u4e14\u9996\u5c3e reads \u8981\u6709\u91cd\u53e0\u3002\u5982\u679c PE 250bp \u6570\u636e\uff0c\u5219 Insert Size \u957f\u5ea6\u8981\u4e3a 400-500 bp\uff08\u9700\u8981\u6ce8\u610f\u7684\u662f\u8f6f\u4ef6\u7684 manual \u4e2d\u53ef\u80fd\u5199\u6210 700bp\uff0c\u8fd9\u662f\u4e0d\u5bf9\u7684<\/a>\uff09\u3002
\n3. \u53ea\u80fd\u4f7f\u7528\u4e00\u4e2a\u6587\u5e93\u7684\u6570\u636e\u3002\uff0c\u4e0d\u652f\u6301\u8f93\u5165 mate paired \u6570\u636e\u3002
\n4. DISCOVAR  de novo (experimental) \u80fd\u8fdb\u884c\u57fa\u56e0\u7ec4\u7684 de novo \u7ec4\u88c5\uff0c\u652f\u6301\u57fa\u56e0\u7ec4\u5927\u5c0f\u53ef\u8fbe ~3 GB\u3002<\/p>\n
3. \u8f6f\u4ef6\u7684\u4f7f\u7528<\/h1>\n
3.1 DISCOVAR \u7684\u4f7f\u7528<\/h2>\n
\u8f6f\u4ef6\u7684\u8f93\u5165\u6587\u4ef6\u662f sort \u8fc7\u540e\u7684 Bam \u6587\u4ef6\uff0c\u4e00\u4e2a\u5e38\u7528\u4f8b\u5b50\uff1a<\/p>\n
\r\n$ Discovar READS=sample-reads.bam REFERENCE=sample-genome.fasta              \\\r\n         REGIONS='10:30892106-30933760' OUT_HEAD=.\/discovar-variants\/assembly\\\r\n         TMP=.\/discovar-variants\/tmp\r\n<\/pre>\n
\u8f6f\u4ef6\u5e38\u7528\u53c2\u6570\uff1a<\/p>\n
\r\nREADS (String)\r\n\u7531\u9017\u53f7\u5206\u5272\u7684\u4e00\u4e9b bam \u6587\u4ef6\uff0c\u6216\u5185\u5bb9\u4e3a\u6bcf\u884c\u4e00\u4e2abam\u6587\u4ef6\u8def\u5f84\u7684 list \u6587\u4ef6\u3002\r\nREGIONS \uff08String\uff09\r\n\u5bf9\u6307\u5b9a\u533a\u57df\u8fdb\u884c\u5206\u6790\u3002\u591a\u4e2a\u533a\u57df\u5219\u7528\u9017\u53f7\u5206\u5272\u3002\u533a\u57df\u7684\u5199\u6cd5\u4e3a chr:start-sotp\u3002\u5982\u679c REGIONS=all\uff0c\u5219\u5bf9\u6240\u6709\u533a\u57df\u8fdb\u884c\u5206\u6790\u3002\r\nTMP \uff08String\uff09\r\n\u6307\u5b9a\u4e34\u65f6\u6587\u4ef6\u8def\u5f84\r\nOUT_HEAD (String)\r\n\u8f93\u51fa\u6587\u4ef6\u7684\u524d\u7f00\u8def\u5f84\r\nNUM_THREADS \uff08unsigned int) default: 0\r\n\u4f7f\u7528\u7684\u7ebf\u7a0b\u6570\u3002\r\nREFERENCE \uff08String)\r\n\u53c2\u8003\u5e8f\u5217 fasta \u6587\u4ef6\u3002\u82e5\u63d0\u4f9b\u6b64\u6587\u4ef6\uff0c\u5219\u80fd\u8fdb\u884c variant calling\uff0c\u5e76\u7ed9\u51fa VCF \u6587\u4ef6\u3002\r\n<\/pre>\n
3.2 DISCOVAR de novo (experimental) \u7684\u4f7f\u7528<\/h2>\n
\u8f6f\u4ef6\u7684\u8f93\u5165\u6587\u4ef6\u662f sort \u8fc7\u540e\u7684 Bam \u6587\u4ef6\u3002\u7a0b\u5e8f\u5728\u8fd0\u884c\u7684\u65f6\u5019\u4f1a\u4f7f\u7528\u6700\u5927\u7684\u7ebf\u7a0b\u6570\u8fdb\u884c\u8fd0\u7b97\u3002<\/p>\n
\r\n$ DiscovarExp --help special\r\n\u4e0a\u8ff0\u547d\u4ee4\u7528\u6765\u67e5\u770b\u8f6f\u4ef6\u7684\u8be6\u7ec6\u53c2\u6570\u3002\r\n$ DiscovarExp READS=sample-reads.bam OUT_DIR=discovarexpOut\r\n\u4e0a\u8ff0\u662f\u8f6f\u4ef6\u7684\u5e38\u7528\u547d\u4ee4\u3002\u540c\u65f6\uff0c\u8f6f\u4ef6\u7684\u53c2\u6570\u975e\u5e38\u5c11\u3002\r\n$ ls discovarexpOut\/a.final\/a.lines.fasta\r\n\u67e5\u770b\u4e3b\u8981\u7ed3\u679c\u3002\r\n<\/pre>\n
4. DISCOVAR\u7ed3\u679c<\/h1>\n
4.1 \u7ed3\u679c\u8868\u73b0\u5f62\u5f0f<\/h2>\n
\"Understanding
\n\u56fe\u4e2d\uff0c\u6bcf\u4e2a\u5355\u72ec\u7684\u7bad\u5934\u79f0\u4e3a edge\uff0c\u8fd9\u4e9b edges \u4ee3\u8868\u7740\u5e8f\u5217\uff1b\u4ece\u8d77\u70b9\u5230\u7ec8\u70b9\uff0c\u6709\u5f88\u591a\u79cd\u4e0d\u540c\u7684\u8def\u5f84\uff0c\u79f0\u4e4b\u4e3a lines\uff1b\u4e0a\u56fe\u4e2d\u6709 4 \u4e2a cells\uff0c\u5176\u4e2d 3 \u4e2a cells \u6709 2 \u4e2a paths\uff0c\u6709 1 \u4e2a cell \u6709 3 \u4e2a paths\u3002
\n\u8fd9\u79cd multiple paths \u53ef\u80fd\u8868\u793a\uff1a\u6742\u5408\u4f4d\u70b9\uff1b\u67d3\u8272\u4f53\u53d8\u5f02\uff1b\u96be\u4ee5\u6d4b\u5e8f\u7684\u4f4d\u70b9\u7b49\u3002<\/p>\n
4.2 DISCOVAR \u7ed3\u679c\u6587\u4ef6 <\/h2>\n
\u751f\u6210\u7684\u7ed3\u679c\u6587\u4ef6\u4f4d\u4e8e discovar-variants\/ \u6587\u4ef6\u5939\u4e0b\uff0c\u4e3b\u8981\u7684\u7ed3\u679c\u6587\u4ef6\u662f\uff1a<\/p>\n
\r\nassembly.final.fasta \u6240\u6709\u7684 edges \u5e8f\u5217 (edges overlap by K-1 bases)\r\nassembly.final.fasta0 \u6240\u6709\u7684 edges \u5e8f\u5217 (without overlaps)\r\nassembly.final.dot dot\u683c\u5f0f\u7684\u7ec4\u88c5\u56fe\r\nassembly.final.variant VCF\u7ed3\u679c\u6587\u4ef6\r\n<\/pre>\n
4.3 DISCOVAR de novo \u7ed3\u679c\u6587\u4ef6<\/h2>\n
\u751f\u6210\u7684\u7ed3\u679c\u6587\u4ef6\u4f4d\u4e8e discovarexpOut\/a.final\/ \u6587\u4ef6\u5939\u4e0b\uff0c\u4e3b\u8981\u7ed3\u679c\u6587\u4ef6\u6709\uff1a<\/p>\n
\r\na.lines.fasta \u591a\u4e2a paths \u4e2d\u4ec5\u9009\u62e9\u7b2c\u4e00\u4e2a path\uff0c\u5f97\u5230\u7684 lines \u5e8f\u5217\u7684 fasta \u6587\u4ef6\u3002\r\na.lines.efasta \u6807\u51c6\u7684 efasta \u6587\u4ef6\uff0c\u6709\u6240\u6709\u7684 paths \u7ed3\u679c\u3002\r\na.fasta \u6240\u6709\u7684 edges \u5e8f\u5217\r\na.lines \u4e8c\u8fdb\u5236\u6587\u4ef6\r\na.lines.src \u4e0a\u4e00\u4e2a\u6587\u4ef6\u7684\u6587\u672c\u5f62\u5f0f\u7ed3\u679c\r\n<\/pre>\n
5. \u603b\u7ed3<\/h1>\n
Discovar \u80fd\u6839\u636e Illumina \u6d4b\u5e8f\u6570\u636e\u6bd4\u5bf9\u5230\u57fa\u56e0\u7ec4\u4e0a\u7684\u7ed3\u679c\u6765\u8fdb\u884c\u57fa\u56e0\u7ec4 de novo \u7ec4\u88c5\uff0c\u5f97\u5230 edges \u5e8f\u5217\uff1b\u82e5\u5728\u63d0\u4f9b\u4e86\u57fa\u56e0\u7ec4\u5e8f\u5217\u7684\u60c5\u51b5\u4e0b\uff0c\u8fd8\u80fd\u8fdb\u884c Vaiants calling\u3002
\nDiscovar de novo (experimental) \u80fd\u6839\u636e Illumina \u6d4b\u5e8f\u6570\u636e\u6bd4\u5bf9\u5230\u57fa\u56e0\u7ec4\u4e0a\u7684\u7ed3\u679c\u6765\u8fdb\u884c\u57fa\u56e0\u7ec4 de novo \u7ec4\u88c5\uff0c\u5f97\u5230 edges \u5e8f\u5217\u3002\u76f8\u6bd4\u4e0e\u524d\u8005\uff0c\u8fd8\u80fd\u5f97\u5230 lines \u5e8f\u5217\uff0c\u8fd9\u662f\u6bd4\u8f83\u5b8c\u6574\u7684\u5e8f\u5217\u6587\u4ef6\u3002<\/p>\n","protected":false},"excerpt":{"rendered":"
1. DISCOVAR\u7b80\u4ecb DISCOVAR \u662f\u6709 ALLPATHS-LG \u8f6f\u4ef6 … \u7ee7\u7eed\u9605\u8bfb →<\/span><\/a><\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":[],"categories":[1],"tags":[],"_links":{"self":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/2252"}],"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=2252"}],"version-history":[{"count":1,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/2252\/revisions"}],"predecessor-version":[{"id":2253,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=\/wp\/v2\/posts\/2252\/revisions\/2253"}],"wp:attachment":[{"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=2252"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=2252"},{"taxonomy":"post_tag","embeddable":true,"href":"http:\/\/www.chenlianfu.com\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=2252"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}