The coverage of intergenic regions is so high in the processing of bacterial RNA-seq reads mapping to reference

Question

[tanxiaojuan1111]

Hi All,

I have two questions about mapping coverage issues to ask you guys. 1) One strain RNAseq reads mapping to reference showed that there is an average of 64% coverage on gene mapping, and remain reads on intergenic mapping. BTW, the stain has complete genome. 2) The other strain RNAseq reads mapping to reference showed that there just is an average 45% coverage on gene mapping, and remain reads on intergenic mapping. However, the reference genome of this strain was sequenced with one PacBio SMRT cell sequencing, and then assembled. The genome was annotated with RAST server.

My questions are :i) why both mapping coverages are so low and ii) how to improve the mapping coverage? Thanks!

[h.mon]

What is the total percentage of reads mapping (gene + intergenic)? Is it above 85%?

Maybe the problem is your annotation, not the mapping. Or the RNA extraction is not being properly cleaned from DNA contaminants.

[tanxiaojuan1111]

Thanks! The first the average of total mapping is greater than 98%. For the second strain, there are 12 samples for RNA-seq project, an average of 9 samples of total mapping is greater than 99%, but an average of remain 3 samples of total mapping just is 77%. I don't know whether these data can be used for analysis on differential expression genes. Look forward to your help! Thanks again!