PASA<\/a>, acronym for P<\/span><\/strong>rogram to A<\/span><\/strong>ssemble S<\/span><\/strong>pliced A<\/span><\/strong>lignments, is a eukaryotic genome annotation tool that exploits spliced alignments of expressed transcript sequences to automatically model gene structures, and to maintain gene structure annotation consistent with the most recently available experimental sequence data. PASA also identifies and classifies all splicing variations supported by the transcript alignments.<\/p>\n Note:<\/strong> \u521b\u5efa\u53ea\u8bfb\u6743\u9650\u7528\u6237\u548c\u6240\u6709\u6743\u9650\u7528\u6237\u5404\u4e00\u4e2a\u3002<\/p>\n \u5b89\u88c5GD\u9700\u8981\u5148\u884c\u5b89\u88c5libgd<\/a><\/p>\n \u518d\u5b89\u88c5GD<\/p>\n \u5b89\u88c5GD\u7684\u76ee\u7684\u662f\u80fd\u901a\u8fc7\u7f51\u9875\u6765\u67e5\u770bPASA\u7684\u8fd0\u884c\u7ed3\u679c\u3002<\/p>\n 2.8.1. \u4fee\u6539PASA\u7684\u914d\u7f6e\u6587\u4ef6$PASAHOME\/pasa_conf\/conf.txt<\/p>\n 2.8.2. \u8be5\u6587\u4ef6\u9700\u8981\u4fee\u6539\u7684\u5730\u65b9\uff1a<\/p>\n 2.8.3. \u4fee\u6539httpd\u914d\u7f6e\u6587\u4ef6\uff0c<\/p>\n \u5728\/etc\/httpd\/conf\/httpd.conf\u6dfb\u52a0\u5982\u4e0b\u51e0\u884c:<\/p>\n \u4e0b\u8f7d\u4e24\u4e2a\u6c61\u67d3\u6570\u636e\u5e93\uff0c\u4e3afasta\u6587\u4ef6\u3002<\/p>\n UniVec_Core includes only oligonucleotides and vectors consisting of bacterial, phage, viral, yeast or synthetic sequences. Vectors that include sequences of mammalian origin are excluded.<\/p>\n PASA\u7684\u4e3b\u7a0b\u5e8f\u662f\uff1a $PASAHOME\/scripts\/Launch_PASA_pipeline.pl, \u5176\u4f7f\u7528\u53c2\u6570\u5982\u4e0b\uff1a<\/p>\n 1. PASA\u7b80\u4ecb PASA, acronym for Program to A … \u7ee7\u7eed\u9605\u8bfb
\nCombine genome and Trinity de novo RNA-Seq assemblies to generate a comprehensive transcript database.<\/p>\n2. PASA\u4f7f\u7528\u524d\u7684\u51c6\u5907<\/h1>\n
2.1 Mysql\u6570\u636e\u5e93\u7684\u51c6\u5907<\/h2>\n
mysql> GRANT SELECT ON *.* TO 'pasa'@'%' IDENTIFIED BY '123456';\r\nmysql> GRANT ALL ON *.* TO 'chenlianfu'@'%' IDENTIFIED BY '123456';\r\nmysql> FLUSH PRIVILEGES;<\/pre>\n
2.1 \u5b89\u88c5perl\u6a21\u5757<\/h2>\n
# cpan\r\ncpan[1]> install DBD::mysql\r\ncpan[1]> install GD<\/pre>\n
2.3 \u5b89\u88c5GMAP<\/a><\/h2>\n
$ wget http:\/\/research-pub.gene.com\/gmap\/src\/gmap-gsnap-2013-03-31.v5.tar.gz\r\n$ tar zxvf gmap-gsnap-2013-03-31.v5.tar.gz\r\n$ cd gmap-2013-03-31\r\n$ .\/configure --prefix=$PWD\r\n$ make -j 8\r\n$ make install<\/pre>\n
2.4 \u5b89\u88c5BLAT<\/a><\/h2>\n
$ wget http:\/\/hgwdev.cse.ucsc.edu\/~kent\/src\/blatSrc35.zip\r\n$ unzip blatSrc35.zip\r\n$ cd blatSrc\r\n$ MACHTYP=x86_64\r\n$ export MACHTYPE\r\n$ mkdir -p ~\/bin\/x86_64\r\n$ make -j 8<\/pre>\n
2.5 \u5b89\u88c5FASTA<\/a><\/h2>\n
$ wget http:\/\/faculty.virginia.edu\/wrpearson\/fasta\/fasta3\/CURRENT.tar.gz\r\n$ tar zxvf CURRENT.tar.gz\r\n$ cd fasta-35.4.12\r\n$ cd src\r\n$ make -f ..\/make\/Makefile.linux_sse2 all\r\n$ cd ..\r\n$ ln -s $PWD\/bin\/fasta35 ~\/bin\/fasta<\/pre>\n
2.6 \u5b89\u88c5PASA<\/h2>\n
$ wget http:\/\/kaz.dl.sourceforge.net\/project\/pasa\/PASA2-r20130425beta.tgz\r\n$ tar zxvf PASA2-r20130425beta.tgz\r\n$ cd PASA2-r20130425beta\/\r\n$ make -j 8<\/pre>\n
2.7 \u5b89\u88c5GD<\/h2>\n
$ wget https:\/\/bitbucket.org\/libgd\/gd-libgd\/get\/93368566388c.zip\r\n$ unzip 93368566388c.zip\r\n$ cd libgd-gd-libgd-93368566388c\r\n$ .\/bootstrap.sh\r\n$ .\/configure\r\n$ make -j 8\r\n$ sudo make install\r\n$ gdlib-config<\/pre>\n
$ wget http:\/\/search.cpan.org\/CPAN\/authors\/id\/L\/LD\/LDS\/GD-2.49.tar.gz\r\n$ tar zxvf GD-2.49.tar.gz\r\n$ cd GD-2.49\r\n$ perl Makefile.PL\r\n$ make -j 8\r\n$ sudo make install<\/pre>\n
2.8 \u914d\u7f6ePASA<\/h2>\n
$ cp $PASAHOME\/pasa_conf\/pasa.CONFIG.template $PASAHOME\/pasa_conf\/conf.txt\r\n$ vim $PASAHOME\/pasa_conf\/conf.txt<\/pre>\n
PASA_ADMIN_EMAIL=(your email address)\r\nMYSQLSERVER=(your mysql server name) \u6b64\u5904\u4e0d\u80fd\u586b\u5199IP\u3002\r\nMYSQL_RO_USER=(mysql read-only username)\r\nMYSQL_RO_PASSWORD=(mysql read-only password)\r\nMYSQL_RW_USER=(mysql all privileges username)\r\nMYSQL_RW_PASSWORD=(mysql all privileges password)\r\nBASE_PASA_URL=http:\/\/server_name\/pasa\/cgi-bin\/<\/pre>\n
# vim \/etc\/httpd\/conf\/httpd.conf\r\n# \/etc\/init.d\/httpd restart<\/pre>\n
ScriptAlias \/pasa \"$PASAHOME\"\r\n<Directory \"$PASAHOME\">\r\n Options MultiViews ExecCGI\r\n AllowOverride None\r\n Order allow,deny\r\n Allow from all\r\n<\/Directory><\/pre>\n
2.9 cleaning the transcript sequences[Optional, requires seqclean to be installed<\/h2>\n
$ cd $PASAHOME\/seqclean\r\n$ tar zxf seqclean.tar.gz\r\n$ cd seqclean\r\n$ wget ftp:\/\/ftp.ncbi.nih.gov\/pub\/UniVec\/UniVec -O UniVec.fasta\r\n$ wget ftp:\/\/ftp.ncbi.nih.gov\/pub\/UniVec\/UniVec_Core -O UniVec_Core.fasta<\/pre>\n
3. PASA\u4e3b\u7a0b\u5e8f\u7684\u4f7f\u7528<\/h1>\n
*\u4ee3\u8868\u8be5\u53c2\u6570\u662f\u5fc5\u987b\u7684<\/strong>\r\n\r\n-c <filename> *<\/span>\r\n\u6bd4\u5bf9\u914d\u7f6e\u6587\u4ef6\u3002\u53ef\u4ee5\u5c06$PASAHOME\/pasa_conf\/pasa.alignAssembly.Template.\r\ntxt\u590d\u5236\u8fc7\u6765\uff0c\u53ea\u662f\u5c06\u5176\u4e2d\u7684MYSQLDB\u4fee\u6539\u6210\u9700\u8981\u7684mysql\u6570\u636e\u5e93\u540d\u3002\r\n\r\n####################\r\n\r\nspliced alignment settings:<\/strong>\r\n--ALIGNERS <string><\/span>\r\n\u6bd4\u5bf9\u7684\u8f6f\u4ef6\uff0c\u53ef\u7528\u7684\u8f6f\u4ef6\u6709gmap\u548cblat\u3002\u4e5f\u53ef\u4ee5\u540c\u65f6\u9009\u62e9\u4f7f\u7528'gmap,blat'\r\n\r\n-N <int> default: 1\r\nmax number of top scoring alignments\r\n\r\n--MAX_INTRON_LENGTH | -I <int> default: 100000<\/span>\r\nmax intron length parameter passed to GMAP or BLAT\r\n\r\n--IMPORT_CUSTOM_ALIGNMENTS_GFF3 <filename><\/span>\r\nonly using the alignments supplied in the corresponding GFF3 file.\r\n\r\n--cufflinks_gtf <filename><\/span>\r\nincorporate cufflinks-generated transcripts\r\n\r\n####################\r\n\r\nactions<\/strong>\r\n-C<\/span>\r\n flag, create MYSQL database\r\n-R<\/span>\r\n flag, run alignment\/assembly pipeline.\r\n-A<\/span>\r\n compare to annotated genes.\r\n--ALT_SPLICE<\/span>\r\n flag, run alternative splicing analysis\r\n\r\n-R \u7528\u4e8e\u6bd4\u5bf9transcripts \uff0c -A \u7528\u4e8e\u548c\u5df2\u6709gff3\u6ce8\u91ca\u6587\u4ef6\u7684\u6bd4\u8f83\u548c\u66f4\u65b0\uff1b\u8fd9\u4e24\u4e2a\u53c2\u6570\u4e0d\r\n\u80fd\u540c\u65f6\u5171\u7528\uff0c\u4f7f\u7528\u4e0d\u540c\u7684\u53c2\u6570\uff0c\u5219 -C \u53c2\u6570\u8bbe\u7f6e\u4e0d\u540c\u7684\u53c2\u6570\u6587\u4ef6\u3002\r\n\r\n####################\r\n\r\ninput files<\/strong>\r\n\r\n-g <filename> *<\/span>\r\n genome sequence FASTA file\r\n\r\n-t <filename> *<\/span>\r\n transcript db\r\n\r\n-f <filename><\/span>\r\n file containing a list of fl-cdna accessions.\r\n\r\n--TDN <filename><\/span>\r\n file containing a list of accessions corresponding to Trinity\r\n (full) de novo assemblies (not genome-guided)\r\n\r\n####################\r\n\r\npolyAdenylation site identification ** highly recommended **<\/strong>\r\n-T<\/span>\r\n flag,transcript db were trimmed using the TGI seqclean tool.\r\n-u <filename><\/span>\r\n value, transcript db containing untrimmed sequences (input to \r\nseqclean).a filename with a .cln extension should also exist, gen\r\nerated by seqclean.\r\n\r\n####################\r\n\r\nJump-starting or prematurely terminating<\/strong>\r\n-x\r\n flag, print cmds only, don't process anything. (useful to get \r\nindices for -x or -e opts below)\r\n-s <int>\r\n pipeline index to start running at (avoid rerunning searches).\r\n-e <int>\r\n pipeline index where to stop running, and do not execute this \r\nentry. \r\n\r\n####################\r\n\r\nMisc:<\/strong>\r\n--TRANSDECODER<\/span>\r\n flag, run transdecoder to identify candidate full-length coding\r\n transcripts\r\n--CPU <int> default: 2<\/span>\r\n multithreading\r\n-d flag, Debug \r\n-h flag, print this option menu and quit<\/pre>\n","protected":false},"excerpt":{"rendered":"