Cufflinks Fpkm

Question: Cufflinks Fpkm

7.7 years ago by

lishiyong • 50 wrote:

Hi: I gain the SOLiD sequencing data.I used bowtie to map human genome then I sort the sam file .I used cuffinks to calculate FPKM with the sam file ,human gtf file .it gives 0 FPKM values and this is for all genes .what's the reason? (1) bowtie -C human_hg18_color -f F3.csfasta -Q F3_QV.qual -v 2 -k 100 -p 6 --mapq --sam test.sam (2) samtools view -uS test.sam 2>/dev/null | samtools sort -m 2000000000 - test.bam (3) cufflinks -G refGene_hg18.gtf test.bam.bam 2011-03-31 lishiyong

cufflinks samtools bam alignment bowtie rna-seq • 2.6k views

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modified 7.7 years ago by Jeremy Goecks • 2.2k • written 7.7 years ago by lishiyong • 50

7.7 years ago by

Che, Anney (NIH/NCI) [E] • 10

Che, Anney (NIH/NCI) [E] • 10 wrote:

Hi lishiyong, Most likely your refGene_hg18.gtf file is not sorted correctly. You have to sort by chr and then by start coordinate. Anney ________________________________________ To: galaxy-user Subject: [galaxy-user] cufflinks FPKM Hi: I gain the SOLiD sequencing data.I used bowtie to map human genome then I sort the sam file .I used cuffinks to calculate FPKM with the sam file ,human gtf file .it gives 0 FPKM values and this is for all genes .what's the reason? (1) bowtie -C human_hg18_color -f F3.csfasta -Q F3_QV.qual -v 2 -k 100 -p 6 --mapq --sam test.sam (2) samtools view -uS test.sam 2>/dev/null | samtools sort -m 2000000000 - test.bam (3) cufflinks -G refGene_hg18.gtf test.bam.bam 2011-03-31 ________________________________ lishiyong

ADD COMMENT • link written 7.7 years ago by Che, Anney (NIH/NCI) [E] • 10

7.7 years ago by

Jeremy Goecks • 2.2k

Jeremy Goecks • 2.2k wrote:

Hello, It's not clear whether you're using Galaxy. If you're using Galaxy, please share you history with me (History Options --> Share/Publish --> Share with User --> my email) and I'll take a look; otherwise, Cufflinks has an email list for questions: tophat.cufflinks@gmail.com Best, J.

ADD COMMENT • link written 7.7 years ago by Jeremy Goecks • 2.2k

7.7 years ago by

Jeremy Goecks • 2.2k

Jeremy Goecks • 2.2k wrote:

If Cufflinks works without a reference GTF, then the problem is the mismatch b/t your GTF and your bam file. A couple things to check: (1) that your genome chrom names in F3.csfata match those in your GTF; if not, you'll need to modify your GTF to match the names in your fasta. (2) that your GTF is sorted as your BAM is sorted. If these issues don't solve your problem, it's best to use the Cufflinks help email that I noted in my previous email. Good luck, J.

ADD COMMENT • link written 7.7 years ago by Jeremy Goecks • 2.2k

7.7 years ago by

Paul Korir • 20

Paul Korir • 20 wrote:

Hi Li, I think the solution lies in changing the chromosome names in the GTF file (refGene_hg18.gtf) from a number e.g. '1' to 'chr1'. Paul 2011/3/31 lishiyong <lishiyong@genomics.org.cn> -- Paul Korir www.paulkorir.com

ADD COMMENT • link written 7.7 years ago by Paul Korir • 20

7.7 years ago by

lishiyong • 50

lishiyong • 50 wrote:

the refgene.gtf have chr in the first colum. and I also sorted it. 2011-03-31 lishiyong 发件人： vasu punj 发送时间： 2011-03-31 22:26:36 收件人： lishiyong; Paul Korir 抄送： galaxy-user 主题： Re: [galaxy-user] cufflinks FPKM the refrence GTF file from Ensembl should have chr in the colum specifying Chrromosome number. Vasu Subject: Re: [galaxy-user] cufflinks FPKM To: "lishiyong" <lishiyong@genomics.org.cn> Cc: "galaxy-user" <galaxy-user@lists.bx.psu.edu> Date: Thursday, March 31, 2011, 8:39 AM Hi Li, I think the solution lies in changing the chromosome names in the GTF file (refGene_hg18.gtf) from a number e.g. '1' to 'chr1'. Paul 2011/3/31 lishiyong <lishiyong@genomics.org.cn> Hi: I gain the SOLiD sequencing data.I used bowtie to map human genome then I sort the sam file .I used cuffinks to calculate FPKM with the sam file ,human gtf file .it gives 0 FPKM values and this is for all genes .what's the reason? (1) bowtie -C human_hg18_color -f F3.csfasta -Q F3_QV.qual -v 2 -k 100 -p 6 --mapq --sam test.sam (2) samtools view -uS test.sam 2>/dev/null | samtools sort -m 2000000000 - test.bam (3) cufflinks -G refGene_hg18.gtf test.bam.bam 2011-03-31 lishiyong ___________________________________________________________ The Galaxy User list should be used for the discussion of Galaxy analysis and other features on the public server at usegalaxy.org. Please keep all replies on the list by using "reply all" in your mail client. For discussion of local Galaxy instances and the Galaxy source code, please use the Galaxy Development list: http://lists.bx.psu.edu/listinfo/galaxy-dev To manage your subscriptions to this and other Galaxy lists, please use the interface at: http://lists.bx.psu.edu/ -- Paul Korir www.paulkorir.com ___________________________________________________________ The Galaxy User list should be used for the discussion of Galaxy analysis and other features on the public server at usegalaxy.org. Please keep all replies on the list by using "reply all" in your mail client. For discussion of local Galaxy instances and the Galaxy source code, please use the Galaxy Development list: http://lists.bx.psu.edu/listinfo/galaxy-dev To manage your subscriptions to this and other Galaxy lists, please use the interface at: http://lists.bx.psu.edu/

ADD COMMENT • link written 7.7 years ago by lishiyong • 50

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