地高辛标记核酸探针斑点杂交法检测轮状病毒疫苗中残余MA－104细胞DNA含量的研究

Application of the Digoxigenin Labelled DNA Probe in Detection of the MA 104 Cell Residual DNA in the Rotavirus Reassortant Vaccine

本文报导用地高辛标记核酸探针和分子斑点杂交技术检测轮状病毒基因重配株Ｌ－３株活疫苗中残余ＭＡ－１０４细胞ＤＮＡ含量的方法。提取和纯化ＭＡ－１０４细胞ＤＮＡ，将其ＡｌｕＩ酶切片段用随机引物法引导ＤＮＡ标记为探针。待检样品抽提核酸后点膜进行斑点杂交。此法灵敏度高，可检出０．１４ｐｇ的ＤＮＡ，特异性强，与非同源性ＤＮＡ无杂交。用此法检测轮状病毒基因重配株Ｌ－３株制备的疫苗，ＭＡ－１０４细胞残余ＤＮＡ含量低于１４ｐｇ低于ＷＨＯ限量标准，结果表明此重配株用于研制疫苗是安全的。

The MA 104 cell DNA probe was labelled with digoxigenin dUTP by method of random primers.The probe was high sensitive and specific,and used to detect the content of MA 104 cell residual DNA in rotavirus reassortant vaccine.It was revealed that the content of residual DNA was less than 100pg per dose.This method of dot blot hybridization with the Dig DNA probe can be used as a routine test for detection of residual DNA in biologicals.