State Key Laboratory of Crop Biology, Agronomy College, Shandong Agricultural University, Tai'an, Shandong Province 271018, P, R, China.
Plant Methods. 2012 Aug 15;8(1):32. doi: 10.1186/1746-4811-8-32.
Multiplex PCR has been successfully applied in many areas since it was first reported in 1988; however, it suffers from poor universality.
A novel method called Universal Multiplex PCR (UM-PCR) was created, which simultaneously amplifies multiple target fragments from genomic DNA. The method has two steps. First, the universal adapter-F and universal adapter-R are connected to the forward primers and the reverse primers, respectively. Hairpin structures and cross dimers of five pairs of adapter-primers are detected. Second, UM-PCR amplification is implemented using a novel PCR procedure termed "Two Rounds Mode" (three and 28-32 cycles). The first round (the first three cycles) is named the "One by One Annealing Round". The second round (28-32 cycles) combines annealing with extension. In the first two cycles of the first round, primers only amplify the specific templates; there are no templates for the universal adapters. The templates of universal adapters begin to be synthesized from the second cycle of the first round, and universal adapters and primers commence full amplification from the third cycle of the first round.
UM-PCR greatly improves the universality of multiplex PCR. UM-PCR could rapidly detect the genetic purity of maize seeds. In addition, it could be applied in other areas, such as analysis of polymorphisms, quantitative assays and identifications of species.
自 1988 年首次报道以来，多重 PCR 已成功应用于许多领域；然而，它的通用性较差。
创建了一种称为通用多重 PCR（UM-PCR）的新方法，该方法可以从基因组 DNA 中同时扩增多个靶片段。该方法有两个步骤。首先，将通用接头-F 和通用接头-R 分别连接到正向引物和反向引物上。检测到五对接头-引物的发夹结构和交叉二聚体。其次，使用一种称为“两轮模式”（三个和 28-32 个循环）的新型 PCR 程序进行 UM-PCR 扩增。第一轮（前三个循环）称为“逐个退火循环”。第二轮（28-32 个循环）结合退火和延伸。在第一轮的前两个循环中，引物仅扩增特定模板；没有通用接头的模板。通用接头的模板从第一轮的第二个循环开始合成，从第一轮的第三个循环开始，通用接头和引物开始完全扩增。
UM-PCR 极大地提高了多重 PCR 的通用性。UM-PCR 可以快速检测玉米种子的遗传纯度。此外，它还可以应用于其他领域，如多态性分析、定量测定和物种鉴定。
