10.3389/fgene.2019.00653.s008 Nishal Kumar Pinna Nishal Kumar Pinna Anirban Dutta Anirban Dutta Mohammed Monzoorul Haque Mohammed Monzoorul Haque Sharmila S. Mande Sharmila S. Mande Table_4_Can Targeting Non-Contiguous V-Regions With Paired-End Sequencing Improve 16S rRNA-Based Taxonomic Resolution of Microbiomes?: An In Silico Evaluation.xlsx Frontiers 2019 metagenomics 16S paired-end sequencing taxonomic profiling microbiome analysis amplicon sequencing 2019-07-12 12:35:33 Dataset https://frontiersin.figshare.com/articles/dataset/Table_4_Can_Targeting_Non-Contiguous_V-Regions_With_Paired-End_Sequencing_Improve_16S_rRNA-Based_Taxonomic_Resolution_of_Microbiomes_An_In_Silico_Evaluation_xlsx/8864159 <p>Background: Next-generation sequencing (NGS) technologies have enabled probing of microbial diversity in different environmental niches with unprecedented sequencing depth. However, due to read-length limitations of popular NGS technologies, 16S amplicon sequencing-based microbiome studies rely on targeting short stretches of the 16S rRNA gene encompassing a selection of variable (V) regions. In most cases, such a short stretch constitutes a single V-region or a couple of V-regions placed adjacent to each other on the 16S rRNA gene. Given that different V-regions have different resolving ability with respect to various taxonomic groups, selecting the optimal V-region (or a combination thereof) remains a challenge.</p><p>Methods: The accuracy of taxonomic profiles generated from sequences encompassing 1) individual V-regions, 2) adjacent V-regions, and 3) pairs of non-contiguous V-regions were assessed and compared. Subsequently, the discriminating capability of different V-regions with respect to different taxonomic lineages was assessed. The possibility of using paired-end sequencing protocols to target combinations of non-adjacent V-regions was finally evaluated with respect to the utility of such an experimental design in providing improved taxonomic resolution.</p><p>Results: Extensive validation with simulated microbiome datasets mimicking different environmental and host-associated microbiome samples suggest that targeting certain combinations of non-contiguously placed V-regions might yield better taxonomic classification accuracy compared to conventional 16S amplicon sequencing targets. This work also puts forward a novel in silico combinatorial strategy that enables creation of consensus taxonomic profiles from experiments targeting multiple pair-wise combinations of V-regions to improve accuracy in taxonomic classification.</p><p>Conclusion: The study suggests that targeting non-contiguous V-regions with paired-end sequencing can improve 16S rRNA–based taxonomic resolution of microbiomes. Furthermore, employing the novel in silico combinatorial strategy can improve taxonomic classification without any significant additional experimental costs and/or efforts. The empirical observations obtained can potentially serve as a guideline for future 16S microbiome studies, and facilitate researchers in choosing the optimal combination of V-regions for a specific experiment/sampled environment.</p>