荧光定量PCR测定粗品肝素钠DNA提取液中反刍基因及猪基因的浓度  

Determination of Concentration of Ruminant Gene and Porcine Gene in DNA Extract of Crude Heparin Sodium by Fluorescence Quantitative PCR

作　　者：李宁[1] 李丽红[1] 米文强 张雪霞[1] 任风芝[1] 刘建芬 LI Ning;LI Lihong;MI Wenqiang;ZHANG Xuexia;REN Fengzhi;LIU Jianfen(New Drug R&D Co.,Ltd.of NCPC,National Engineering Research Center of Microbial Medicine,Hebei Industry Microbial Metabolic Engineering & Technology Research Center,Shijiazhuang 050015,China;Huakun Hebei Biological Technology Co.,Ltd.of NCPC,Shijiazhuang 050000,China)

机构地区：[1]华北制药集团新药研究开发有限责任公司,微生物药物国家工程研究中心,河北省工业微生物药物代谢工程技术中心,河北石家庄050015 [2]华北制药华坤河北生物技术有限公司,河北石家庄050000

出　　处：《化学与生物工程》2021年第1期65-68,共4页Chemistry & Bioengineering

摘　　要：采用细胞裂解和蛋白酶K消化技术,结合DNA制备柱选择性吸附DNA的方法提取粗品肝素钠中的动物DNA,将DNA标准溶液、粗品肝素钠DNA提取液分别与相应的PCR试剂混合,进行PCR扩增,测定粗品肝素钠DNA提取液中反刍基因及猪基因浓度。结果表明,反刍基因浓度在0.01~1000 pg·μL-1范围内线性关系良好,猪基因浓度在0.1~10000 pg·μL-1范围内线性关系良好。该方法具有良好的专属性、准确度、精密度,适用于粗品肝素钠动物来源的种属鉴别。We extracted the animal DNA in crude heparin sodium by cell lysis and protease K digestion,combined with selective adsorption of DNA by DNA preparation column.Moreover,we mixed the DNA standard solution and the DNA extract of crude heparin sodium with the corresponding PCR reagents,respectively,for PCR amplification,and determined the concentrations of ruminant gene and porcine gene in the DNA extract of crude heparin sodium.The results show that the concentration of ruminant gene shows a good linearity in the range of 0.01-1000 pg·L-1,and the concentration of porcine gene shows a good linearity in the range of 0.1-10000 pg·L-1.The method has good specificity,accuracy,and precision,which is suitable for the species identification of animal origin of crude heparin sodium.

关 键 词：粗品肝素钠 荧光定量PCR 种源鉴别 

分 类 号：R446.1[医药卫生—诊断学]