酶标探针在转基因植物检测中的应用
华中科技大学生命科学与技术学院中英联合实验室, 分子生物物理学教育部重点实验室, 武汉 430074
收稿日期: 2007-04-26
修回日期: 2007-07-08
网络出版日期: 2007-12-10
Application of enzyme-labeled probe in testing of transgenic plant
Received date: 2007-04-26
Revised date: 2007-07-08
Online published: 2007-12-10
采用碱性磷酸酶标记DNA制备分子探针, 并首次在植物中应用。酶在苯醌作用下与单链DNA联结, 形成DNA和酶的共价复合物即酶标探针。此探针通过分子杂交与待测DNA结合, 再与酶的底物作用显色, 3~6h 内可观察结果。用此探针检测转基因植物中的UidA基因, 点杂交和Southern杂交结果表明, 所合成的酶标探针具有快速、准确、安全而经济的优点。点杂交证明外源UidA基因被成功转化到受体植物中, Southern 杂交对转基因的材料检测的结果证明, 该材料包含多个外源UidA基因拷贝, 初步确定其外源UidA基因拷贝数在5个以上。
涂知明,陈泠,杨广笑,何光源 . 酶标探针在转基因植物检测中的应用[J]. 遗传, 2007 , 29(12) : 1533 -1537 . DOI: 10.1360/yc-007-1533
Used alkaline-phosphatase-labeled DNA as a probe to examine the expression of foreign UidA gene in transgenic plants. Alkaline phosphatase coupled with polyethyleneimine (PEI) using P-benzoquine as the cross-linking reagent was covalently linked to single-stranded DNA via glutraldehyde. Such DNA-enzyme complexes were used as a probe for dot hybridization and Southern blot. After hybridization and incubation with a substrate solution, results can be observed directly in three to six hours and the results showed that it was a sensitive, specific, rapid, safe and economical probe. Dot hybridization analysis showed that the UidA gene was transformed into the target plants and southern blot showed that there were at least 5 copies of UidA gene in transgenic plants.
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