CAJ | 학술논문

报道了一种粘性末端的限制性酶切片段的直接克隆和测序的方法.对限制性酶切片段的粘性末端先用T4 DNA聚合酶处理,变为平末端,然后用TaqTM DNA聚合酶在其3′末端加上A腺苷,即可利用T/A克隆载体进行直接克隆测序.利用这种简单而快速的方法,对2个RFLP探针Psr680的限制性酶切片段(1.65 kb和0.65 kb)进行了测序,表明这种方法可以替代利用相应载体进行相应酶切等处理的粘性末端连接克隆测序的方法.
보도료일충점성말단적한제성매절편단적직접극륭화측서적방법.대한제성매절편단적점성말단선용T4 DNA취합매처리,변위평말단,연후용TaqTM DNA취합매재기3′말단가상A선감,즉가이용T/A극륭재체진행직접극륭측서.이용저충간단이쾌속적방법,대2개RFLP탐침Psr680적한제성매절편단(1.65 kb화0.65 kb)진행료측서,표명저충방법가이체대이용상응재체진행상응매절등처리적점성말단련접극륭측서적방법.
A method is presented by which DNA restriction fragments with cohesive-ends are enzymatically blunt-ended with T4 DNA polymerase and then a single 3′ A-adenosine is added by TaqTM DNA polymerase before ligating directly into a T/A cloning vector.Using this rapid and simple method,two restriction fragments (1.65 kb and 650 bp) from a RFLP probe Psr680 were directly cloned into a T/A cloning vector,and then were sequenced.This method is an alternative method to cohesive-end ligation of restricted fragments,which uses a prepared vector containing the corresponding restrictive enzyme site.