Importing 3 Casava 1.8 files

[Tym (Tymofii Sokolskyi)]

Hi! I am very new to Qiime2, and I have received 16S MiSeq data that I need to analyze. I received 3 files from the sequencing center - titled 5730-P1_S1_L001_I1_001, 5730-P1_S1_L001_R1_001 and 5730-P1_S1_L001_R2_001 so I assumed it is in Casava 1.8 format however I am still confused as to the files I got - I haven’t found topics on the forums showing how to correctly import using I1, R1 and R2.

I tried importing the R1 and R2 through Casava 1.8, viewed the quality plot, selected the trimming length and ran dada2, but I received an error message
An error was encountered while running DADA2 in R (return code -9), please inspect stdout and stderr to learn more.

I did it because I thought my data was already demultiplexed by the center but then I noticed that demux.qzv showed just 1 sample while there should be 45. Here is the quality plot. So this means that my data is multiplexed, right? Then how should I proceed?

Thank you and Merry Christmas!

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[thermokarst]

Hi @Tym!

@Tym

so I assumed it is in Casava

Actually, it looks like you have multiplexed EMP-style reads (Casava 1.8 is a demultiplexed format):

https://docs.qiime2.org/2019.10/tutorials/importing/#emp-protocol-multiplexed-paired-end-fastq

Take a look at the link above and let us know how it goes!

[Tym (Tymofii Sokolskyi)]

Thank you! But how do I extract barcodes, forward and reverse reads from those files?

[elximo (Hazem Sharaf)]

I just had a similar problem and followed the instructions on the protocol.

I found that I needed to rename the files first after gzip compression as follows: “forward” (for R1), “reverse” (R1) and “barcodes” (for I) with all of them having the fastq.gz extension.

After that import them as EMPPairedEndSequences and then demultiplex them.

[Tym (Tymofii Sokolskyi)]

Thanks! I actually tried that but got an error for demultiplexing. And I also thought that R1 and R2 files contain both forward and reverse reads each just for two runs?

[thermokarst]

@Tym

I actually tried that but got an error for demultiplexing

Please share the commands run (copy and paste), and the complete error observed (copy and paste when run with the --verbose flag).

@Tym

And I also thought that R1 and R2 files contain both forward and reverse reads each just for two runs?

Can you explain this question in a bit more detail? R1 and R2 are convention (usually) for forward (R1) and reverse (R2), but it isn't a rule.