Question: Low reads mapping in methylation sequencing
0
gravatar for sjbasu
3 months ago by
sjbasu0
India
sjbasu0 wrote:

Hello People,

I have four methylation samples (MeDIP-seq) of two related plant species, testing methylation in case of saline stress. details are as follows:

Sample 1: Species 1 no stress

Sample 2: Species 1 with saline stress

Sample 3: Species 2 no stress

Sample 4: Species 2 with saline stress

Now when I map these reads to the genome (using BWA) I get 95 % mapping for Sample 1 and 3 while Sample 2 and 4 shows 23 % mapping. I am not able to understand the reason. All samples have ~25 mill reads and the sample extraction and quality of reads really is good !! Has anyone faced this problem ?? What can be the cause ??

mapping medip • 173 views
ADD COMMENTlink modified 3 months ago by Chris Miller18k • written 3 months ago by sjbasu0
0
gravatar for Chris Miller
3 months ago by
Chris Miller18k
Washington University in St. Louis, MO
Chris Miller18k wrote:

Pull them up in IGV - did you get read-through into the adapters? Then adapter trimming might help. You might also try using a methylation aware aligner (BSmap, Biscuit, etc)/

ADD COMMENTlink written 3 months ago by Chris Miller18k

Hello Chris,

I followed your advice and used Bismark (in Bowtie1 mode) and the mapping percentage dropped even more at 3%.

Is it that the methylated-Cytosines (C) are converted to thymine (T) in the methylated DNA extracted and hence the sequenced read is also having T whereas in reference genome all these points with T are actually C ??!!

Any idea what can be going wrong ??!!!

ADD REPLYlink written 3 months ago by sjbasu0
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