Yajima R
Research Group, R/D, Dainabot Co., Ltd., Diagnostics Division, Matsudo.
Rinsho Byori. 1997 Jan;45(1):41-6.
There are several methods that have been developed for the detection of a small amount of nucleic acids. Ligase chain reaction(LCR) is a highly sensitive assay for the detection of a specific DNA sequence. LCR utilizes four oligodeoxynucleotides(probes) complementary to each of target DNA strands and repeats a thermalcycling amplification step. Two pairs of oligodeoxynucleotides renature adjacent to one another on each of the separated target DNA strands, resulting in nick formation. A thermostable DNA ligase joins the nick covalently. Each ligated product can serve as a template sequence in subsequent rounds of thermalcycling (denaturation, annealing/ligation). In this manner LCR accumulates the ligated products exponentially by repeating thermalcycling. One of the distinctive features of LCR is to be able to detect a known point-mutation easily. Gap-LCR, a modified LCR, has been developed to improve the sensitivity and the specificity of the standard LCR by setting up gaps (3 < or = nucleotides) at the ligatable 3' and 5' termini of the probes. The gap-LCR probes form a short gap after annealing of the probes to the target DNA. The gap is filled in by a thermostable DNA polymerase and the resultant nick is sealed by a thermostable DNA ligase to generate the ligated probes. Gap-LCR repeats this step on thermalcycling and accumulates the LCR products in an exponential fashion.
已经开发出几种用于检测少量核酸的方法。连接酶链反应（LCR）是一种用于检测特定DNA序列的高度灵敏的检测方法。LCR利用与每条目标DNA链互补的四种寡脱氧核苷酸（探针），并重复热循环扩增步骤。两对寡脱氧核苷酸在每条分离的目标DNA链上彼此相邻复性，导致切口形成。热稳定的DNA连接酶将切口共价连接。每个连接产物可在随后的热循环（变性、退火/连接）轮次中用作模板序列。通过这种方式，LCR通过重复热循环以指数方式积累连接产物。LCR的一个显著特点是能够轻松检测已知的点突变。间隙LCR是一种改良的LCR，通过在探针可连接的3'和5'末端设置间隙（3个或更多核苷酸）来提高标准LCR的灵敏度和特异性。间隙LCR探针与目标DNA退火后形成一个短间隙。该间隙由热稳定的DNA聚合酶填充，产生的切口由热稳定的DNA连接酶封闭以生成连接的探针。间隙LCR在热循环中重复此步骤，并以指数方式积累LCR产物。
