Comparison of 16S rRNA Gene Based Microbial Profiling Using Five Next-Generation Sequencers and Various Primers

• DOI: 10.3389/fmicb.2021.715500
• Publication Date: 2021-10-14T09:39:40Z
• Authors: Changwoo Park; Seung Bum Kim; Sang Ho Choi; Seil Kim
• Keywords: Minion

Microbial community analysis based on the 16S rRNA-gene is used to investigate both beneficial and harmful microorganisms in various fields environments. Recently, next-generation sequencing (NGS) technology has enabled rapid accurate microbial analysis. Despite these advantages of NGS metagenomics study, sample transport, storage conditions, amplification, library preparation kits, sequencing, bioinformatics procedures can bias results. In this eight mock communities were pooled from genomic DNA Lactobacillus acidophilus KCTC 3164T, Limosilactobacillus fermentum 3112T, gasseri 3163T, Lacticaseibacillus paracasei subsp. 3510T, reuteri 3594T, Lactococcus lactis 3769T, Bifidobacterium animalis 5854T, breve 3220T. The DNAs quantified by droplet digital PCR (ddPCR) mixed as communities. amplified with rRNA gene universal primer pairs sequenced MiSeq, IonTorrent, MGIseq-2000, Sequel II, MinION platforms. a comparison primer-dependent bias, profiles V1-V2 V3 regions similar original ratio communities, while V1-V3 region relatively biased. platform-dependent sequence read short-read platforms (MiSeq, MGIseq-2000) showed lower than that long-read (Sequel II MinION). Meanwhile, sequences biased some data all regions, L. was greatly underrepresented generally overrepresented. samples index (BI) calculated PCA performed for comparison. relative abundance high BI values separated particular, rich AT GC poses problems genome assembly, which lead bias. According comparative analysis, development reference material (RM) been proposed calibrate microbiome