常规PCR法快速鉴别不同种属来源的粗品肝素钠  

Rapid identification of crude heparin sodium from various species origins by routine PCR

作　　者：陈川[1] 朱思宇[1] 李丽[1] 胡淑媛[1] 罗都强[1,2] 

机构地区：[1]河北大学生命科学学院,河北保定071002 [2]河北大学药物化学与分子诊断省部共建教育部重点实验室,河北保定071002

出　　处：《中国生物制品学杂志》2014年第5期705-708,共4页Chinese Journal of Biologicals

基　　金：国家自然科学基金(3117188;30901755);教育部长江学者创新团队发展计划(IRT1124)

摘　　要：目的采用常规PCR法快速鉴别不同种属来源的粗品肝素钠。方法通过柱回收和胶回收两步回收法,从粗品肝素钠中提取动物残留核酸,以其为模板,通过常规PCR法进行种属来源的鉴定。分别以猪、羊来源的核酸为模板,用3种种属特异性引物(猪、羊1、羊2)进行PCR扩增,验证引物的特异性;对柱回收、胶回收和两步回收法提取的核酸样品及含不同比例(1%、5%、10%、15%)羊来源肝素钠的混合样品进行PCR检测;并检测3批次肝素钠样品的种属来源。结果猪的引物能特异性扩增猪来源的核酸,羊的引物能特异性扩增羊来源的核酸;采用单一胶回收和柱回收法提取的残留核酸,不能有效地利用常规PCR鉴定种属来源,而两步回收法提取的残留核酸,能有效地利用常规PCR鉴定种属来源;采用两步回收法提取核酸,可检测到含1%羊来源肝素钠的混合样品;3批次的肝素钠样品中均混有羊肝素钠。结论常规PCR法操作简便迅速,成本较低,可用于不同种属来源的粗品肝素钠的定性检测。Objective To identify the crude heparin sodium from various species origins by routine PCR. Methods Residual animal nucleic acids were extracted from crude heparin sodium by two step（column and gel）recovery method and used as templates for identification of species origin by routine PCR. The specificities of three primers were verified by PCR using the DNAs from pig and sheep origins as templates. The nucleic acid samples extracted by column recovery, gel recovery and two step recovery as well as mixed samples containing various proportions（1%,5%,10% and 15%）of heparin sodium were determined by PCR. Meanwhile,the species origins of heparin sodium of various batches were tested. Results The nucleic acids from pig origin were amplified specifically by using the pig-specific primer,while those from sheep origin by the two sheep-specific primers. The species origins of residual nucleic acids extracted by gel recovery or column recovery alone could not be identified effectively,while those extracted by two step recovery could be identified effectively,by routine PCR. The residual nucleic acid in mixed samples containing 1% heparin sodium from sheep origin extracted by two step recovery was detected by PCR. The heparin sodium from sheep origin was detected in the heparin sodium samples of three batches. Conclusion Routine PCR method was simple,rapid and low-cost,which might be used for qualitative determination of crude heparin sodium from various species origins.

关 键 词：肝素钠 聚合酶链反应 种属 

分 类 号：Q789[生物学—分子生物学]