HE Lihong~(1; 2) YU Rongli~1 CHEN Mingjie~(1*) PAN Yingjie~3(~1Key Laboratory of Applied Mycological Resources and Utilization; Ministry of Agriculture; the People’s Republic of China; Shanghai Research Center for Edible Fungi Biotechnology and Engineering; Shanghai Key Laboratory of Agricultural Genetics and Breeding; Institute of Edible Fungi; Shanghai Academy of Agricultural Sciences; Shanghai 201106; China; ~2Department of Microbiology; College of Life Sciences; Nanjing Agricultural University; Nanjing; Jiangsu 210095; China; ~3Shanghai Ocean University; Shanghai 200090; China)
【中文摘要】 <正>An efficient and cost effective method for obtaining good quality DNA from composts containing high levels of organic matter has been developed.The protocol consisted of sample washing with sodium phosphate and EDTA,cell wall disruption using combined chemical and enzyme treatment prior to extraction,removal of humic acid and other inhibitors in high salt buffer containing polyvinyl polypyrrolidone (PVPP),and precipitation of DNA with polyethylene glycol (PEG-8000).The method was used t...更多o extract 16S ribosomal DNA from four samples taken at different stages in the preparation of PhaseⅡcompost used for Agaricus bisporus cultivation.Extracted DNA was sufficiently free of contaminants to allow PCR amplification of products derived from bacteria,actinomycetes and fungi.Purified products derived from PCR amplification of the extracted 16S rDNA using primers F27 and R1498 were used to construct 16S rDNA libraries.PCR amplification of ten randomly selected positive clones using primer M13 revealed that the 16S rDNA insertion rate was higher than 90%. 还原
【英文摘要】 An efficient and cost effective method for obtaining good quality DNA from composts containing high levels of organic matter has been developed.The protocol consisted of sample washing with sodium phosphate and EDTA,cell wall disruption using combined chemical and enzyme treatment prior to extraction,removal of humic acid and other inhibitors in high salt buffer containing polyvinyl polypyrrolidone (PVPP),and precipitation of DNA with polyethylene glycol (PEG-8000).The method was used to extract 16S ribosomal DNA from four samples taken at different stages in the preparation of PhaseⅡcompost used for Agaricus bisporus cultivation.Extracted DNA was sufficiently free of contaminants to allow PCR amplification of products derived from bacteria,actinomycetes and fungi.Purified products derived from PCR amplification of the extracted 16S rDNA using primers F27 and R1498 were used to construct 16S rDNA libraries.PCR amplification of ten randomly selected positive clones using primer M13 reve...更多aled that the 16S rDNA insertion rate was higher than 90%. 还原
【英文关键词】 Agaricus bisporus; compost phaseⅡ; bacterial 16S rDNA library
【基金】Supported by project of China Agricultural Ministry (No.nyhyzx-07-008)
【文献出处】 食用菌学报,Acta Edulis Fungi,编辑部邮箱,2008年04期 【DOI】CNKI:SUN:SYJB.0.2008-04-007