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CN105274214A 测定人PON1基因rs662位点多态性的方法及试剂盒 无效 - 驳回

测定人PON1基因rs662位点多态性的方法及试剂盒
摘要：
测定人PON1基因rs662位点多态性的方法及试剂盒。该方法包括：提供待测人基因组DNA；提供扩增人PON1基因rs662位点附近序列的上游引物和下游引物，其中下游引物具有错配碱基A；以所述待测人基因组DNA为模板，利用上游引物和下游引物进行PCR扩增反应以获得含RATTC片段或RATT片段的扩增产物，其中R为人PON1基因rs662位点上的待定碱基A或G；提供限制性内切酶；利用限制性内切酶对所得扩增产物进行酶切以获得相应的酶切产物；以及根据所得酶切产物来确定人PON1基因rs662位点上的待定碱基R是A还是G。限制性内切酶为仅能够切开GATTC片段与AATTC片段之一，或者GATT片段与AATT片段之一的限制性内切酶。本发明的测定手段不但快速可靠，而且测定成本大大降低。
摘要（英）：

The method relates to a method and a kit for determining the rs662 site polymorphism of a human PON1 gene. The method comprises the following steps: providing a human genome DNA to be determined; providing an upstream primer and a downstream primer for amplifying a sequence nearby the rs662 site of the human PON1 gene, wherein the downstream primer has a mismatching base A; carrying out a PCR amplification reaction by using the upstream primer and the downstream primer with the human genome DNA to be determined as a template to obtain an amplification product containing an RATTC fragment or an RATT fragment, wherein R is an undetermined base A or G on the rs662 site of the human PON1 gene; providing a restrictive endonuclease; carrying out enzyme digestion on the amplification product by using the restrictive endonuclease to obtain a corresponding enzyme digestion product; and determining the undetermined base R on the rs662 site of the human PON1 gene is A or G according to the obtained enzyme digestion product. The restrictive endonuclease is a restrictive endonuclease which only can cut one of a GATTC fragment and an AATTC fragment or one of a GATT fragment and an AATT fragment. The determination method is fast and reliable, and greatly reduces the determination cost.

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