Question: Chip-Seq's mapping rate problem.(too low)
0
gravatar for leenaehyeon
3.8 years ago by
leenaehyeon10
Korea, Republic Of
leenaehyeon10 wrote:

Chip-Seq's mapping rate problem.

I've runned FastQC and result is not bad.

 

but mapping rate is horrible (read tophat_out/allign_summary.txt)

Left reads:
          Input     :  14775968
           Mapped   :    133948 ( 0.9% of input)
            of these:     36875 (27.5%) have multiple alignments (1612 have >20)
Right reads:
          Input     :  14775968
           Mapped   :    123596 ( 0.8% of input)
            of these:     30111 (24.4%) have multiple alignments (1430 have >20)
 0.9% overall read mapping rate.

Aligned pairs:     91752
     of these:     15121 (16.5%) have multiple alignments
                    6000 ( 6.5%) are discordant alignments
 0.6% concordant pair alignment rate.

 

First, using bbmap --> mapping(tophat2)

I don't know why...

 

chip-seq • 2.0k views
ADD COMMENTlink written 3.8 years ago by leenaehyeon10
1

If it's Chip-Seq why did you use Tophat? Although I don't think this is the problem. Try to run FastQC after bbmap

ADD REPLYlink written 3.8 years ago by Asaf6.1k

I usually using tophat2 for RNA-seq, so I tried same way for Chip-seq.

Is there another way for Chip-seq analysis tool?

ADD REPLYlink written 3.8 years ago by leenaehyeon10

Bowtie/Bowtie2 is normally used for ChIP-seq.

http://bowtie-bio.sourceforge.net/index.shtml 

It runs within Tophat so it will already be installed. It also comes with ready-made genome libraries. Maybe try aligning against some pre-made libraries as well just in case something went wrong with your genome build.

ADD REPLYlink written 3.8 years ago by jotan1.2k
1

I'm really sorry for this question but I have to ask. Are you sure you're working on the right genome? I've accidentally tried to align mouse data to human genome and vice versa.

ADD REPLYlink written 3.8 years ago by jotan1.2k

My boss said that genome download from UCSC.. :(((( ..hmm..

I'll try another way.. Thank you for reply!

ADD REPLYlink written 3.8 years ago by leenaehyeon10

#[1] bbmap
/home/Downloads/bbmap/bbduk.sh in=A1.fastq in2=A2.fastq out=A1_bbduk.fastq out2=A2_bbduk.fastq ref=/home/Downloads/bbmap/resources/truseq.fa.gz k=13 ktrim=r qtrim=t trimq=10 minlength=20

#[2] mapping

tophat2 -o tophat_out_A -G /home/Documents/genome/mm10/mm10.gtf --transcriptome-index /home/Documents/genome/mm10/mm10_transcripts --library-type fr-firststrand /home/Documents/genome/mm10/mm10 A1_bbduk.fastq A2_bbduk.fastq

 

Is that wrong?

ADD REPLYlink written 3.8 years ago by leenaehyeon10
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