I'm working with two different batches of samples and I want to compare them. First batch of samples used V3-V4 regions (primers 341F 785R) and the second one used V4 region (primers 515F 806R). Thus, I know that they have some similar region and I can compare them. However, I would need to trunc the first ones or to merge the second ones and compare to the reverse sequence of the firsts... Something to just work with the same region...
Someonw know how can I do it??
Really really thanks in advance!!
For the case in which you have data from amplicon region, but with different primer pairs, there are a few options you can try below. Note: regardless of the approach you use, you'll still have to worry about slight variations in PCR / sequencing biases in your results. Which may inflate differences between data sets. It only takes a single base in a primer to alter what you observe in your data.
- Alter the trim & truncation settings for each run, using these slightly different primers, such that the the same ASVs will be generated. Then you can merge the tables and sequence files for down stream processing.
Closed reference OTU picking is another option. I suggest you read He et. al. 2015, Rideout et. al. 2014, Wescot et. al. 2015, and finally Callahan et. al. 2017 for more information.
Just as an FYI, I know other mods have previously combined data sets with different V3V4 primer sets. In at least one case, only the reverse primer is different between the two. Thus, they used the 1st approach mentioned above, as they only had to trim a few bases of the end of one of the sequencing runs.
Thanks @SoilRotifer for the headstart.
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