A POLYMERASE CHAIN-REACTION MEDIATED BY A SINGLE PRIMER - CLONING OF GENOMIC SEQUENCES ADJACENT TO A SEROTONIN RECEPTOR PROTEIN CODING REGION
A POLYMERASE CHAIN-REACTION MEDIATED BY A SINGLE PRIMER - CLONING OF GENOMIC SEQUENCES ADJACENT TO A SEROTONIN RECEPTOR PROTEIN CODING REGION
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DOI：
10.1093/nar/19.25.7155
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发表时间：
1991-12-01
影响因子：
14.9
通讯作者：
SHENK, T
中科院分区：
文献类型：
--
作者：
PARKS, CL;CHANG, LS;SHENK, T
在适当的条件下，基因组DNA序列经聚合酶链式反应扩增后可产生特异的双链DNA产物，该聚合酶链式反应只包含一个单一的引物。这种类型的扩增反应是用各种引物和底物DNA进行的。除了非特异性的异质产物外，11个引物中的5个重复地指导双链DNA的合成，该双链DNA对应于模板中包含真正的引物退火点的区域。克隆了其中3个扩增产物，并对其末端进行了测序。所有三个都在5‘端各含有一份引物，其中一条的位置代表了真实的引物结合部位。在每种情况下，第二个拷贝的引物的位置表明它最初与模板DNA中的部分同源序列杂交。这一单引物反应使扩增和克隆与已知DNA序列相邻的未知序列的DNA区域成为可能。这里描述的一个单引物反应产物包括5-羟色胺受体基因编码区5‘端的序列，该序列包含一个功能性启动子。
Under appropriate conditions, specific double-stranded DNA product was generated after amplification of genomic DNA sequences in a polymerase chain-like reaction that contained only a single primer. This type of amplification reaction was performed with a variety of primers and substrate DNAs. In addition to nonspecific heterogeneous products, 5 of 11 primers reproducibly directed synthesis of double-stranded DNA that corresponded to the region of the template that contained the authentic primer annealing site. Three of these amplified products were cloned and their ends were sequenced. All three contained a copy of the primer at both 5' ends, and the position of one of the primers represented the authentic primer binding site. In each case, the location of the second copy of the primer indicated that it had initially hybridized to a partially homologous sequence in the template DNA. This single primer reaction makes it possible to amplify and clone a DNA region of unknown sequence that is adjacent to a known DNA sequence. One of the single primer reaction products described here included sequence to the 5' side of the coding region of a serotonin receptor gene that contained a functional promoter.