荧光定量PCR检测猪肝中猪链球菌2型的基因组DNA提取方法的比较
Comparison of Genome DNA Extraction Methods of Streptococcus suis Type 2 from Pig Liver by Fluorescent Quantitative PCR Detection
凌淑萍 1赵健 1陈国 1叶宇飞 1许秀琴 1吕燕 1吴银良1
作者信息
- 1. 宁波市农业科学研究院,浙江宁波315000
- 折叠
摘要
[目的]在人工布菌猪肝中评估溶菌酶-苯酚法和柱式纯化试剂盒法提取猪链球菌2型基因组DNA在荧光定量检测中的应用效果,为猪链球菌2型的荧光定量检测选择合理的基因组DNA提取方法提供试验依据.[方法]采用溶菌酶-苯酚法和柱式纯化试剂盒法分别提取人工布菌猪肝样本中猪链球菌2型基因组DNA,通过荧光定量PCR技术检测猪链球菌2型的相对含量.[结果]利用荧光定量方法能够检测到猪链球菌2型.该方法可以最低检测到12 CFU/g的猪链球菌,特异性良好,标准曲线方程为y=-3.185 9x+39.901 0.在人工布菌猪肝样本中,溶菌酶-苯酚法在qPCR实验中检测猪链球菌2型较灵敏.[结论]溶菌酶-苯酚法在猪肝样品中提取基因组DNA效果较好;试剂盒法在细菌纯培养物中提取基因组DNA具有安全、高效等优点.
Abstract
[Objective] To evaluate the difference between lysozyme-phenol method and purification kit method in extracting the Streptococcus suis type 2 from pig liver,and to provide experimental basis for selecting an appropriate genomic DNA extraction method in the application of fluorescent quantitative detection of Streptococcus suis type 2.[Method] Both lysozyme-phenol method and purification kit method were used to extract the genome DNA of Streptococcus suis type 2 from pig liver samples.The relative contents of Streptococcus suis type 2 were detected by fluorescent quantitative PCR technology.[Result] Fluorescent quantitative detection could detect the Streptococcus suis type 2.This method could detect at least 12 CFU/g Streptococcus suis,with good specificity.The equation of standard curve was y =-3.185 9x +39.901.Lysozyme-phenol method was more sensitive in qPCR test on the detection of Streptococcus suis type 2.[Conclusion] Lysozyme-phenol method is better in extracting genomic DNA from pig liver samples.However,purification kit method is widely applied in the fluorescent quantitative detection due to the advantages of safety and efficiency in extraction of genomic DNA.
关键词
猪链球菌2型/基因组DNA/qPCR检测Key words
Genomic DNA/Streptococcus suis type 2/qPCR detection引用本文复制引用
出版年
2016
国家科技期刊平台
NSTL
维普
万方数据