一种无外源菌DNA污染的鸡球虫卵囊的制备方法  

A method for preparing chicken coccidial oocysts without contamination of exogenous bacterial DNA

作　　者：龚振兴[1,2] 刘保红 马雪婷[1,2] 殷昊 李法财[1,2] 蔡建平 GONG Zhenxing;LIU Baohong;MA Xueting;YIN Hao;LI Facai;CAI Jianping(Lanzhou Veterinary Research Institute/Key Laboratory of Veterinary Parasitology of Gansu Province/State Key Laboratory of Veterinary Etiological Biology,Chinese Academy of Agricultural Sciences,Lanzhou 730046,China;Jiangsu Collaborative Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou 225009,China)

机构地区：[1]中国农业科学院兰州兽医研究所/家畜疫病病原生物学国家重点实验室/甘肃省动物寄生虫病重点实验室,兰州730046 [2]江苏省动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《黑龙江畜牧兽医》2020年第5期108-111,151,共5页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(31272554);中国农业科学院农业科技创新工程专项(2014-LVRI-09)。

摘　　要：为了获得无鸡肠道和粪便细菌DNA污染的鸡球虫卵囊,试验采用实验室制备的无微生物污染的水配制相关试剂,对柔嫩艾美耳球虫卵囊采用漂浮纯化、灭菌处理和外源菌DNA去除等方法,获得高纯度的柔嫩艾美耳球虫卵囊;提取卵囊的基因组DNA,用细菌16S rRNA基因通用引物进行PCR扩增并进行序列分析,以鉴定该卵囊是否被外源菌DNA污染。结果表明:提取的柔嫩艾美耳球虫卵囊基因组DNA经PCR方法检测外源DNA呈阴性。说明用此方法制备的柔嫩艾美耳球虫卵囊无外源菌DNA污染,可用于制备高纯度的无外源菌DNA污染的鸡球虫卵囊。In order to obtain chicken coccidia oocysts that without contamination of chicken gut and fecal bacterial DNA,the microbial DNA-free water was used to prepare the related reagents for floating purification,sterilization treatment and exogenous DNA removal of Eimeria tenella oocysts,and high-purity Eimeria tenella oocysts were obtained.The genomic DNA of oocysts was extracted and amplified by PCR by using 16 S RNA gene universal primers of bacteria,and sequence analysis was performed to identify whether the E.tenella oocytes were contaminated by exogenous bacteria DNA.The results showed that the E.tenella oocytes prepared by this method had no DNA pollution of exogenous bacteria,and the method could be used to prepare high-purity chicken coccidial oocysts without DNA pollution of exogenous bacteria.

关 键 词：鸡球虫 卵囊 制备 基因组DNA 外源菌DNA 

分 类 号：S852.723[农业科学—基础兽医学]