Köseoğlu Ozgen, Kocagöz Tanil
Hacettepe Universitesi Tip Fakültesi, Mikrobiyoloji ve Klinik Mikrobiyoloji Anabilim Dali, Ankara.
Mikrobiyol Bul. 2004 Jul;38(3):239-43.
The use of cloning vectors has revolutionized molecular biology. Any vector with appropriate cloning sites can be used to clone a section of DNA and polymerase chain reaction (PCR) is a useful method for producing DNA fragments that are intended to be cloned. When Taq polymerase is used in PCR for polymerization, the enzyme adds an extra adenosyl (A) nucleotide to the 3' end of the extended strand, in a template independent manner. T-cloning vectors are created by adding a thymidine (T) residue to the ends of the cloning site. This aids ligation with PCR products possessing A-T. In this study, we prepared a T-cloning vector system, which guarantees insertion of open reading frames to the right position for expression. This method provides an easy way of cloning PCR products assuring at the same time in frame insertion.
克隆载体的使用彻底改变了分子生物学。任何具有合适克隆位点的载体都可用于克隆一段DNA,而聚合酶链反应(PCR)是产生拟克隆DNA片段的一种有用方法。当在PCR中使用Taq聚合酶进行聚合反应时,该酶会以不依赖模板的方式在延伸链的3'端添加一个额外的腺苷(A)核苷酸。T克隆载体是通过在克隆位点的末端添加一个胸腺嘧啶(T)残基而构建的。这有助于与具有A-T的PCR产物进行连接。在本研究中,我们制备了一种T克隆载体系统,该系统可确保将开放阅读框插入到正确的位置进行表达。此方法提供了一种克隆PCR产物的简便方式,同时确保框内插入。