Trimming/ derep error

[tobaiasbama]

Hi, am running 16s Hifi demultiplexed analysis, after running the last line of code in this chunk:
meta_ana <- "~/Users/oluwatobifijabi/Downloads/Elijah/Metagenomics Results 2"
path.out <- "Figures/"
path.rds <- "RDS/"
fns1 <- list.files(meta_ana, pattern="fastq.gz", full.names=TRUE)
F27 <- "AGRGTTYGATYMTGGCTCAG"
R1492 <- "RGYTACCTTGTTACGACTT"
rc <- dada2:::rc
theme_set(theme_bw())
nops1 <- file.path(meta_ana, "noprimers", basename(fns1))
prim1 <- removePrimers(fns1, nops1, primer.fwd=F27, primer.rev=dada2:::rc(R1492), orient=TRUE) ; i got this
warning message:
In removePrimers(fns1, nops1, primer.fwd = F27, primer.rev = dada2:::rc(R1492), :
No reads passed the primer detection.

[benjjneb]

No reads are passing primer detection. This means that the removePrimers function is not finding any reads that contain both the primers you provided, and thus is not creating an output file, at least for some samples.

Can you say more about what type of data you are working with? Is this amplicon sequencing data? if so, what are the primers being used? What is the sequencing technology? Have you confirmed the primers are on the reads?

[tobaiasbama]

Metagenomics Workflow 16S rRNA (1).docx
here is a link to the data(https://drive.google.com/file/d/1SWwLXrwoilnrKvuZ8-tm62rZcTiStRoj/view?usp=drivesdk)

[benjjneb]

I would start by confirming that the expected primers are actually on the reads you are working with. The identify primers section of the dada2 tutorial for ITS data shows an example of this: https://benjjneb.github.io/dada2/ITS_workflow.html#identify-primers