摘要
本研究旨在分析不同DNA提取方法对半滑舌鳎(Cynoglossus semilaevis)肠道菌群结构分析结果的影响,并了解健康半滑舌鳎肠道菌群结构特征。通过溶菌酶结合超声波裂解法(CLU)、Zirmil-beating细胞破碎方法(ZBC)和QIA商业试剂盒三种方法提取半滑舌鳎肠道菌群基因组DNA,采用Illumina MiSeq高通量测序分析三种方法提取的样品DNA,比较其菌群结构特征和多样性。结果显示,三种方法提取的样品经检测共得到7516个OTU,而且鉴定的优势菌门一致,主要包括变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)、螺旋菌门(Spirochaetae)、拟杆菌门(Bacteroidetes)。通过Alpha多样性指数分析,CLU、ZBC、QIA方法对应的Chao1指数分别为5668、5868、4090;Simpson指数分别为0.021、0.022、0.018,QIA方法的Chao1和Simpson指数均低于其他两种方法。基于UniFrac距离矩阵进行Beta多样性分析显示CLU和QIA方法提取的样品之间的微生物群落结构和物种丰富度更相似。本研究表明不同方法提取的DNA经高通量测序分析鉴定到的主要优势菌群类别一致,但对一些丰度较低的菌群类别鉴定时存在偏差。
The purpose of this study was to analyze the effects of different DNA extraction methods on structural analysis of intestinal microbiota from C.semilaevis based on high throughput sequencing and comprehensively learn about the structural characteristics of its intestinal flora.In this study,three DNA extraction methods including combination of lysozyme and ultrasonic lysis(CLU),Zirmil-beating cell disruption(ZBC)and QIA commercial kits were used to extract the genomic DNA of intestinal microflora from healthy C.semilaevis.The DNA samples were sequenced on an Illumina MiSeq high-throughput sequencing platform,and the structural characteristics and diversities of the intestinal microflora were analyzed.The results showed that a total of 7516 OTUs were obtained across all samples extracted by the three DNA extraction methods.The identified dominant bacterial taxa were similar among these samples.At Phylum level,the dominant bacterial taxa included Proteobacteria,Firmicutes,Spirochaetae and Bacteroides.Through Alpha diversity index analysis,the corresponding Chao1 indexes of CLU,ZBC and QIA methods were 5668,5868 and 4090 respectively,and the Simpson index were 0.021,0.022 and 0.018 respectively.The Chao1 and Simpson indexes of QIA methods were all lower than those of other methods.Beta diversity analysis based on the UniFrac distance matrix showed that the microbial community structure and species richness were more similar between the CLU and QIA samples.These results indicated that these three different DNA extraction methods reached the similar results in the major dominant intestinal microflora,based on the investigation using high-throughput sequencing analysis.However,biases occurred in the identification results of some less abundant intestinal microflora.The results providereference for research methods of the diversity and structure of fish intestinal microflora,especially the DNA extraction strategy,and contribute to comprehensively expounding the structural characteristics of intestinal microflora of C.semilaevis.
作者
葛仑华
江博赟
孙敬锋
吕爱军
胡秀彩
陈丽梅
Ge Lunhua;Jiang Boyun;Sun Jingfeng;LüAijun;Hu Xiucai;Chen Limei(College of Fisheries,Tianjin Agricultural University,Tianjin 300392,China)
出处
《天津农学院学报》
CAS
2022年第3期29-35,共7页
Journal of Tianjin Agricultural University
基金
国家自然科学基金项目(31972840)
天津市自然科学基金项目(19JCZDJC34600)
天津市企业科技特派员项目(18JCTPJC64700,19JCTPJC60100)。
关键词
高通量测序
DNA提取方法
半滑舌鳎
肠道菌群
组成结构
high-throughput sequencing
DNA extraction method
C.semilaevis
intestinal microflora
composition structure
