Hi everyone, I would like to ask why the splicing rate between the left and right ends of my data is only 1%, and what are the possible reasons for this result? (Experimental and raw letter analysis) And how can we improve it?
sequencing region: 16S rRNA gene V4 (515F ,806R)
Looks like you are loosing most of the reads at merging step. That's means that youre reads do not overlap. Solution will be:
- Reconsider truncation parameters (set it to higher values)
- Decrease minoverlap in Dada2 parameters
- If nothing works use only forard or reverse reads for the analysis.
Thank you for your prompt reply, I'll go back and try!
For solution No.3, Which one should I use for subsequent analysis, and will there be a big difference between the forward and reverse reads?
It depends on the quality plots and region targeted. Usually I just go for forward reads. Note that using only one read (F or R) may lead to poorer taxonomy annotations, so I use it only when there is no other way to improve merging stats.
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