## ----filenames, message=FALSE, warning=FALSE------------------------------- library(dada2); packageVersion("dada2") fnF1 <- system.file("extdata", "sam1F.fastq.gz", package="dada2") fnR1 <- system.file("extdata", "sam1R.fastq.gz", package="dada2") filtF1 <- tempfile(fileext=".fastq.gz") filtR1 <- tempfile(fileext=".fastq.gz") ## ----inspect--------------------------------------------------------------- plotQualityProfile(fnF1) # Forward plotQualityProfile(fnR1) # Reverse ## ----filter---------------------------------------------------------------- filterAndTrim(fwd=fnF1, filt=filtF1, rev=fnR1, filt.rev=filtR1, trimLeft=10, truncLen=c(240, 200), maxN=0, maxEE=2, compress=TRUE, verbose=TRUE) ## ----derep----------------------------------------------------------------- derepF1 <- derepFastq(filtF1, verbose=TRUE) derepR1 <- derepFastq(filtR1, verbose=TRUE) ## ----learn, warning=FALSE-------------------------------------------------- errF <- learnErrors(derepF1, multithread=FALSE) # multithreading is available on many functions errR <- learnErrors(derepR1, multithread=FALSE) ## ----dada, warning=FALSE--------------------------------------------------- dadaF1 <- dada(derepF1, err=errF, multithread=FALSE) dadaR1 <- dada(derepR1, err=errR, multithread=FALSE) print(dadaF1) ## ----merge----------------------------------------------------------------- merger1 <- mergePairs(dadaF1, derepF1, dadaR1, derepR1, verbose=TRUE) ## ----bimeras--------------------------------------------------------------- merger1.nochim <- removeBimeraDenovo(merger1, multithread=FALSE, verbose=TRUE) ## ----sample2, warning=FALSE------------------------------------------------ # Assign filenames fnF2 <- system.file("extdata", "sam2F.fastq.gz", package="dada2") fnR2 <- system.file("extdata", "sam2R.fastq.gz", package="dada2") filtF2 <- tempfile(fileext=".fastq.gz") filtR2 <- tempfile(fileext=".fastq.gz") # Filter and Trim filterAndTrim(fwd=fnF2, filt=filtF2, rev=fnR2, filt.rev=filtR2, maxN=0, trimLeft=10, truncLen=c(240, 200), maxEE=2, compress=TRUE, verbose=TRUE) # Dereplicate derepF2 <- derepFastq(filtF2, verbose=TRUE) derepR2 <- derepFastq(filtR2, verbose=TRUE) # Infer sample composition (using already learned error rates) dadaF2 <- dada(derepF2, err=errF, multithread=FALSE) dadaR2 <- dada(derepR2, err=errR, multithread=FALSE) # Merge merger2 <- mergePairs(dadaF2, derepF2, dadaR2, derepR2, verbose=TRUE) ## ----make-table------------------------------------------------------------ seqtab <- makeSequenceTable(list(merger1, merger2)) seqtab.nochim <- removeBimeraDenovo(seqtab, verbose=TRUE) dim(seqtab.nochim)