Method of detecting a polynucleotide sequence and labelled polynucleotides useful therein
申请人:Malcolm, Alan David Blair
公开号:EP0124221A1
公开(公告)日:1984-11-07
A method of detecting nucleic acid sequences is disclosed and enzyme-labelled reagents for use therein. According to the invention sample polynucleotide is first hybridised with a polynucleotide probe comprising a nucleic acid sequence complementary to that which is to be detected, and having a polynucleotide tail, preferably a single stranded poly(dA) or poiy(dT) tail. Following attachment of the probe to the sample, an enzyme-containing marker, is attached to the probe, the marker comprising a polynucleotide having a single-stranded portion, to the base groups of which is or are attached one or more enzymatically active groups, and a polynucleotide tail which can be attached to the probe by annealing or hybridisation. Preferably the marker has a single-stranded polynucleotide tail complementary to that on the probe whereby the marker can be attached to the probe by a second hybridisation step. Presence of the sequence to be detected is confirmed by testing the labelled sample for any enzyme related activity.
本发明公开了一种检测核酸序列的方法及其使用的酶标记试剂。根据本发明,样品多核苷酸首先与多核苷酸探针杂交,探针由与待检测的核酸序列互补的核酸序列组成,具有多核苷酸尾部,最好是单链 poly(dA) 或 poiy(dT) 尾部。将探针附着在样品上后,含酶标记物会附着在探针上,标记物包括具有单链部分的多核苷酸,其碱基组上附着一个或多个酶活性基团,以及可通过退火或杂交附着在探针上的多核苷酸尾部。标记物最好具有与探针上的单链多核苷酸尾部互补的多核苷酸尾部,这样标记物就可以通过第二杂交步骤连接到探针上。通过检测标记样品是否具有与酶相关的活性来确认是否存在要检测的序列。