采样方法对冷鲜鸡表面细菌DNA提取及高通量测序结果的影响  

Effects of Different Sampling Methods on Microbial DNA Extraction from Chilled Chicken and the High-Throughput Sequencing of Amplification Products

作　　者：肖英平[1] 何祥祥[2] 戴宝玲 桂国弘 唐标[1] 杨华[1] 

机构地区：[1]浙江省农业科学院农产品质量标准研究所,浙江省植物有害生物防控省部共建国家重点实验室培育基地,浙江杭州310021 [2]西北农林科技大学食品科学与工程学院,陕西杨凌712100

出　　处：《食品科学》2017年第24期260-264,共5页Food Science

基　　金：浙江省公益技术运用研究项目(2016C32073);浙江省植物有害生物防控重点实验室--省部共建国家重点实验室培育基地项目(2010DS700124-ZM1608)

摘　　要：目的:比较涂抹法和冲洗法对冷鲜鸡表面细菌DNA提取及高通量测序结果的影响。方法:将4只冷鲜鸡对半分开,分别采用涂抹法和冲洗法采集其表面微生物,提取基因组DNA,对细菌16S rRNA基因V3~V4区进行聚合酶链式反应扩增,通过Illumina Hiseq测序平台对扩增产物进行高通量测序,使用QIIME等软件对测序序列进行分析统计。结果:两种方法采集的样品在细菌基因组DNA提取、菌群丰富度、多样性及菌群结构等方面均无显著差异(P>0.05)。高通量测序结果表明,冷鲜鸡表面细菌菌群主要分布于7个门,10个属。变形菌门为优势菌门,占70%以上;希瓦氏菌属、假单胞菌属、不动杆菌属、嗜冷杆菌属和环丝菌属为优势菌属,分别占10%~20%。结论:两种采样方法对冷鲜鸡表面细菌DNA提取及高通量测序没有明显影响,但冲洗法比涂抹法简单易行,且不易发生二次污染,因此更具有实用性。本实验同时也为冷鲜鸡表面细菌的研究提供了数据资料。Objective：To compare the swabbing and rinsing methods used for sampling chilled chicken by evaluating their effects on microbial DNA extraction and subsequent high-throughput sequencing.Methods：Four chilled chickens were divided into equal halves and sampled by superficial swabbing and rinsing methods for each half,respectively.Microbial genomic DNA was extracted from the collected samples and the V3-V4 region of the bacterial 16S rRNA gene was amplified by PCR.The PCR products were then subjected to high-throughput sequencing on an Illumina Hi Seq sequencing platform.The obtained sequences were processed and analyzed using QIIME and other softwares.Results：There was no difference in bacterial genomic DNA extraction or bacterial community richness,diversity and structures between the samples collected by the two methods（P〉0.05）.High-throughput sequencing showed that the bacterial community in chilled chicken samples consisted mainly of 7 phyla and 10 genera.Proteobacteria was the dominant phylum,accounting for more than 70% of the bacterial community;Shewanella,Pseudomonas,Acinetobacter,Psychrobacter,and Brochothrix were the dominant genera,each representing 10%-20% of the bacterial community.Conclusions：The two sampling methods exhibit no obvious difference in bacterial DNA extraction from chilled chicken or high-throughput sequencing.However,compared with swabbing,rinsing has the advantages of easy operation and non-secondary contamination,so it is more practical in microbiological analysis of chilled chicken.The experiment also provides useful data for the study of bacteria on chilled chicken.

关 键 词：冷鲜鸡 菌群结构 涂抹法 冲洗法 高通量测序 

分 类 号：TS251.55[轻工技术与工程—农产品加工及贮藏工程] TS207.4[轻工技术与工程—食品科学与工程]