I recently learned that Deblur works with only forward reads, and if we have forward and reverse reads for sequences, we must join the sequences prior to performing sequence filtering thru Deblur. I am done with joining my sequences. Right before I denoise my sequences with Deblur, I was curious to check the quality of their bases. Attached below is a screenshot of the quality score graph for my joined reads:
The following are the questions that I need help with:
- Can someone help me in understanding if the quality of my joined reads are good to proceed further? This is the first time I am seeing the values take a dip and then rise even further, to be honest. Does this happen quite often?
- The first 30-40 bases have surprisingly low scores, therefore I was thinking of using the "--p-left-trim-len n" parameter of the deblur-16S method to eliminate the presence of the initial few bases. Would that be the right way to approach this issue?
- For bases between 150 and 300, the quality scores are above 40 - is that normal and can these be included while performing the deblur-16S filtering step? In other words, what would be a good --p-trim-length parameter value for me? The tutorial mentions that the Deblur developers recommend 115 to 130, however if I were to choose the same for my sequences, I would be dropping majority of my bases.
Looking forward to hearing your thoughts on this.