Department of Chemical Engineering & Institute of Biotechnology and Chemical Engineering, I-Shou University, Kaohsiung 84008, Taiwan.
IEEE Trans Nanobioscience. 2013 Jun;12(2):119-27. doi: 10.1109/TNB.2013.2258469.
Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) is a commonly used laboratory technique and useful in small-scale basic research studies of complex genetic diseases that are associated with single nucleotide polymorphisms (SNPs). Before PCR-RFLP assay for SNP genotyping can be performed, a feasible primer pair observes numerous constraints and an available restriction enzyme for discriminating a target SNP, are required. The computation of feasible PCR-RFLP primers and find available restriction enzymes simultaneously aim at a target SNP is a challenging problem. Here, we propose an available method which combines the updated core of SNP-RFLPing with a genetic algorithm to reliably mine available restriction enzymes and search for feasible PCR-RFLP primers. We have in silico simulated the method in the SLC6A4 gene under different parameter settings and provided an appropriate parameter setting. The wet laboratory validation showed that it indeed usable in providing the available restriction enzymes and designing feasible primers that fit the common primer constraints. We have provided an easy and kindly interface to assist the researchers designing their PCR-RFLP assay for SNP genotyping. The program is implemented in JAVA and is freely available at http://bio.kuas.edu.tw/ganpd/.
聚合酶链反应-限制片段长度多态性（PCR-RFLP）是一种常用的实验室技术，对于与单核苷酸多态性（SNP）相关的复杂遗传疾病的小规模基础研究很有用。在进行 SNP 基因分型的 PCR-RFLP 检测之前，需要观察到许多可行的引物对的约束条件，并且需要有可用的限制酶来区分目标 SNP。同时计算可行的 PCR-RFLP 引物和寻找可用的限制酶针对目标 SNP 是一个具有挑战性的问题。在这里，我们提出了一种结合 SNP-RFLPing 更新核心和遗传算法的可行方法，以可靠地挖掘可用的限制酶并搜索可行的 PCR-RFLP 引物。我们在不同参数设置下对 SLC6A4 基因进行了计算机模拟，并提供了一个适当的参数设置。湿实验室验证表明，它确实可以用于提供可用的限制酶，并设计符合常见引物约束的可行引物。我们提供了一个简单友好的界面，以帮助研究人员设计他们的 SNP 基因分型的 PCR-RFLP 检测。该程序是用 JAVA 实现的，并可在 http://bio.kuas.edu.tw/ganpd/ 免费获取。
