利用不同方法从深加工牦牛肉产品中提取基因组DNA效果的比较  

Comparison of Quantity and Quality of Genomic DNAs Extracted from Highly Processed Yak Meat Products Using Different Methods

作　　者：何建文[1,2,3] 韩建林[4,2,3] 罗玉柱[1] 

机构地区：[1]甘肃农业大学动物科学与技术学院,兰州730070 [2]中国农业科学院国际家畜研究所畜禽牧草遗传资源联合实验室 [3]中国农业科学院北京畜牧兽医研究所,北京100193 [4]中国农业科学院北京畜牧兽医研究所农业部畜禽遗传资源与利用重点开放实验室,北京100193

出　　处：《生物技术通报》2010年第10期162-167,共6页Biotechnology Bulletin

基　　金：“十一五”国家科技支撑计划项目(2007BAD52B05);CAAS-ILRI联合研究项目(2008-CI-03)

摘　　要：为了从深加工牦牛肉产品中获得片段较大、数量较多的基因组DNA,以用于分子追溯的检测,尝试并系统地比较了异硫氰酸胍法、酚-氯仿抽提法和试剂盒提取法的提取效果。通过定量分析、凝胶电泳和PCR片段大小梯度扩增等手段,对利用3种方法提取的基因组DNA的数量和质量进行了比较。结果显示,利用酚-氯仿抽提法提取的基因组DNA质量不佳,异硫氰酸胍法和试剂盒提取法的提取效果相当,PCR可以扩增出730 bp左右的核外基因细胞色素b,而核内基因只能扩增出300bp左右的核基因DNA片段(内乳铁蛋白基因,271 bp;BoLA-DRB3基因,302 bp)。综合考虑提取时间、费用、数量和质量等各方面的因素,认为异硫氰酸胍法操作简便、快速、廉价、有效,是较好的从动物深加工肉产品中提取基因组DNA的方法。To recover sufficient amount of large fragments of the genomic DNAs from highly processed yak meat products for their molecular traceability tests, this study explored three extraction protocols of phenol-chloroform extraction, guanidine thiocyauate method and commercial DNA extraction kit, separately. The quality and quantity of the extracted DNAs were assessed using three methods of quanti- tative analysis through spectrophotometer,argarose gel electrophoresis and PCR amplifications for different sizes of mtDNA and nuclear genom- ic gene fragments. The results showed that the DNAs extracted using both the guanidine thiocyanate method and DNA extraction kit have bet- ter quality and quantity that enabled the PCR amplifications of both 730 bp fragment of Cyt b gene from the mtDNA and 300 bp nuclear DNA fragments （271 bp of lactoferrin gene; 302 bp of BoLA-DRB3 gene）. However, the DNAs recovered following the phenol-chloroform method failed completely in the amplifications. In consideration of all the factors including time and cost required for and also quantity and quality of the DNAs recovered from different extraction protocols,we suggest that guanidine thioeyanate method is a simple, rapid ,cheap and sufficient protocol that can be used for routine DNA extraction from the highly processed yak meat products for their further molecular traceability tests.

关 键 词：异硫氰酸胍法 酚-氯仿抽提法 试剂盒方法 深加工牦牛肉产品 基因组DNA 

分 类 号：S823[农业科学—畜牧学]