Influence of PCR cycle number on microbial diversity analysis through next generation sequencing.

Yunhe AN; Lijuan Gao; Junbo LI; Yanjie Tian; Jinlong Wang; Xuejuan Zheng; Huijuan WU

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Influence of PCR cycle number on microbial diversity analysis through next generation sequencing.

Author: Yunhe AN ¹ ; Lijuan GAO ¹ ; Junbo LI ¹ ; Yanjie TIAN ¹ ; Jinlong WANG ¹ ; Xuejuan ZHENG ¹ ; Huijuan WU ¹
Author Information

1. Beijing Center For Physical and Chemical Analysis, Beijing 100089, China.
Publication Type:Journal Article
Keywords: PCR cycle number; high throughput sequencing; microbial diversity analysis
MeSH: Animals; Bacteria; classification; DNA, Bacterial; genetics; Gene Library; High-Throughput Nucleotide Sequencing; Polymerase Chain Reaction; methods; RNA, Ribosomal, 16S; genetics; Rumen; microbiology; Sheep; Soil Microbiology
From: Chinese Journal of Biotechnology 2016;32(8):1115-1123
CountryChina
Language:Chinese
Abstract: Using of high throughput sequencing technology to study the microbial diversity in complex samples has become one of the hottest issues in the field of microbial diversity research. In this study, the soil and sheep rumen chyme samples were used to extract DNA, respectively. Then the 25 ng total DNA was used to amplify the 16S rRNA V3 region with 20, 25, 30 PCR cycles, and the final sequencing library was constructed by mixing equal amounts of purified PCR products. Finally, the operational taxonomic unit (OUT) amount, rarefaction curve, microbial number and species were compared through data analysis. It was found that at the same amount of DNA template, the proportion of the community composition was not the best with more numbers of PCR cycle, although the species number was much more. In all, when the PCR cycle number is 25, the number of species and proportion of the community composition were the most optimal both in soil or chyme samples.