Previously, I have asked questions related with mapping the miRNAs. (miRNA mapping rate is very low.. (less than 0.03%))
Thank you David! :) Finally I could successfully map my miRNA reads.
But, this time I had another set of samples..but same design. 3 controls, VS 3 treated..
I followed exactly same logic.. ( since they are generated same machine.)
- Remove adapter sequence
- Remove index sequence
But, This time, I realized that after removing the adapter sequences, (TGGAATTCTCGGGTGCCAAGGAACTCCAGTCAC), I can see the file size are reduced dramatically, which means most of reads are removed.
For example, here is the fastq file size for original files
c1 (265M), c2 (428M), c3 (248 M), a1(268M), a2(344M), a3(443M)
after removing the adapter sequences
c1 (132M, okay), c2( 15M, weird), c3(208M, okay), a1(153M, okay), a2(15M, weird), a3(18M, weird)
When I looked at the fastq files, I can see those files (e.g. c2, a2, a3,), many of reads are mostly composed of adapter sequences (I am not sure why it is... maybe experiments were bad? No idea about experiments). I guess that this is the reason that file are mostly chopped.
When, I try to further analysis (e.g. remove index sequence, first 4, last 4 removal for my case), I ran bowtie2.
Here is the result of bowtie2.
c1 c2 c3 a1 a2 mapping rate 55.88% 7.75% 70.06% 68.14% 27.48% Total number of reads 1196717 135524 1841729 1367558 134217
I am wondering whether I can further process of this analysis. I heard that mapping rate should be usually around 50-80%. In my case, it is much less than that. Also the number of total reads are so less.
I need some comments for this. Is it the problem of experiments? OR what else? Can I further analyze this?