Merged reads lower frequency

User Support
1

Hello,

We are trying to decide whether to use just the forward read or use merged reads for our data. We used PEAR to merge the reads on qiime version 2019.4. Once completed, we discovered that our total frequency dropped as well as number of features when compared to just the forward read. Why would the frequency drop when with the merged reads?

PairedTable.qzv (1007.1 KB)
ForwardTable.qzv (1.1 MB)

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2

Hi @Marinab2,

There are a few reasons you would see the drop. First, it could be the number of reads you pass in - if your merge isn’t great you might lose reads there. Second, there may be a quality issue (hard to know without earlier artifacts). That quality issue is probably worse because DADA2 assumes you’re passing in single ended reads and doesn’t behave properly on already-joined reads.

Best,
Justine

3

Do you suggest running the forward and reverse reads through DADA2 first and then pairing them?

I've attached my demuxed output and dada2 stats.

stats-dada2.qzv (1.2 MB)
per_sample_sequences.qzv (293.6 KB)

4

Hi @Marinab2,

Running paired end denoising in DADA2, it will do the merging for you. Your sequences look high quality and like there should be no issue with denoising - although depending on the length of your hypervariable region, you may not be able to join the sequences.

Best,
Justine

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5

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