Similar protocols

Pipeline publication

[…] is limited to 16S rRNA gene-based identification of C. difficile and cannot predict whether a strain produces toxin or carries a functional pathogenicity locus, consideration for accurate identification of C. difficile and related members may be useful in assessing clinical outcomes of new microbial therapies that rely on 16S rRNA gene sequencing to validate recovery of the microbiota., Whole-genome shotgun sequence datasets available from (i) The Wellcome Trust Sanger Institute and (ii) The University of Maryland Institute for Genome Sciences designated as novel C. difficile isolates were downloaded from the NCBI Sequence Read Archive (see Table  for accessions), trimmed for quality using Trimmomatic and assembled into contigs using Minia. Contigs containing portions of 16S rRNA genes were identified using BLASTN and extracted for amplicon simulations. For each isolate, we subsequently simulated 16S rRNA amplicon sequence reads (10,000 per isolate) from the V4 region (the primary amplicon region selected in the real datasets) with a random nucleotide error rate of 0.5%. The Diagnostic True Positive Rate was computed as the percentage of sequences unambiguously assigned by Resphera Insight to C. difficile., For false positive assessment, simulated V4 sequences were generated from reference 16S rRNA genes for 22 unique species within the Clostridium XI cluster (10,000 per species, 0.5% nucleotide error rate). False positives were defined as unambiguous assignments to C. difficile., Raw 16S rRNA […]

Pipeline specifications

Software tools Trimmomatic, Minia, BLASTN
Databases SRA