不同热处理方法对鲤鱼DNA的影响及不同DNA提取方法的比较  

Effects of heating on DNA of carp and the different methods for DNA extraction

作　　者：雍凌[1] 张文众[1] 刘玉梅[1,2] 贾旭东[1] 严卫星[1] 

机构地区：[1]中国疾病预防控制中心营养与食品安全所,北京100021 [2]北京联合大学应用文理学院,北京100191

出　　处：《中国食品卫生杂志》2011年第3期245-248,共4页Chinese Journal of Food Hygiene

基　　金：转基因生物检测和监测新技术食品中转基因作物溯源体系研究(2008ZX08012-005)

摘　　要：目的研究不同加热方法处理后鲤鱼基因组DNA长度和含量的变化,并对4种DNA提取方法进行比较。方法用电磁炉和微波炉将鲤鱼背部肌肉加热煮沸1、2、3 h取出,再分别用酚氯仿法、Wizard试剂盒法、磁珠法和离心柱法提取处理前后鲤鱼基因组DNA,利用琼脂糖凝胶电泳对处理前后的总DNA长度进行比较,再用实时荧光定量PCR法检测所得DNA样品中内参基因β-actin含量的变化。结果琼脂糖凝胶电泳结果显示,处理后DNA片段明显降解。生鱼肉提取β-actin的量为1 070 213.39拷贝/μl,电磁炉处理1、2、3 h的浓度分别为143 309.6、441 350.43和256 994.16拷贝/μl,微波炉处理1、2、3 h的浓度分别为269 121.17、267 371.16和134 649.97拷贝/μl。酚氯仿法、Wizard试剂盒法、离心柱法和磁珠法提取的平均DNA浓度分别为718 944.93、737 945.64、26 751.22和2 934.06拷贝/μl。结论加热煮沸导致DNA降解,因此加工食品中DNA的检测应当选择较短的目标片段。4种DNA提取方法比较,Wizard试剂盒法和酚氯仿法提取的DNA浓度显著高于离心柱法和磁珠法,且离心柱法又高于磁珠法。Objective To investigate the effects of different heating methods on the degradation of carp DNA and to compare different DNA extraction methods.MethodsThe dorsal muscle of carp was boiled for 1,2 and 3 h by electromagnetic oven or microwave oven.Genomic DNA of carp meat was extracted by four different methods（phenol/chloroform extraction,Wizard commercial Kit,Mag-Bind DNA Kit and spin column）.The length of DNAs before and after heat treatment was compared by agarose gel electrophoresis.The contents of β-actin in tested DNA samples were determined by real-time PCR assays.ResultsThe contents of β-actin DNA after heating for 1,2 and 3 h in electromagnetic oven were 143309.6,441350.43 and 256994.16 copy/μl and those in microwave oven were 269121.17,267371.16 and 134649.97 copy/μl respectively.The contents of raw carp β-actin DNA was 1070213.39 copy/μl,which were decreased significantly after being heated.However,the contents of β-actin DNA would be no significant change with extended heating time.The contents of DNA extracted by phenol/chloroform or Wizard commercial Kit（718944.93 and 737945.64 copy/μl respectively） were higher than that by spin column and Mag-Bind DNA Kit（26751.22 and 2934.06 copy/μl respectively）.ConclusionDNA could be degraded by heating for more than 1 h.Therefore,the target fragments of DNA in food selected for DNA detection should be shorter.As for the extraction methods,the contents of DNA extracted by phenol/chloroform and Wizard commercial Kit were higher than that by spin column and Mag-Bind DNA Kit.

关 键 词：加热时间 鲤鱼DNA DNA提取方法 荧光定量聚合酶链反应 

分 类 号：Q78[生物学—分子生物学]