I found that the raw paired sequences were mixed, there should only have the sequences guided by 515F primer in the raw-r1.fq, and there should only have the sequences guided by 806R primer in the raw-r2.fq, respectively.
The sequences was obtained from illumina Miseq PE300.
I asked the sequencing company for details but they cannot explain the reason of this mixed situation.
So I wonder if anyone have found this problem too, and please tell me the possible reason and solutions. Thank you very much.