CYP450热点突变基因芯片分型方法的建立

Genotyping of hot-spot CYP450 SNPs with DNA microarray

目的：探索适合多位点同步检测的SNP扩增方案和杂交检测条件，建立芯片杂交信号判读的标准。方法：设计多重扩增的引物和探针；优化荧光标记引物的多重PCR条件。自行制备氨基化片基并点样制片，激光共聚焦扫描后判读检测结果。结果：野生与突变探针杂交信号比值大于4或小于2．5对分型结果的判断有意义，而当信号值在2．5～4范围内应重新检测分析。结论：通过多片段同步扩增和不同位点平衡杂交的实现，本方法适于流行病学调查，体现了寡核苷酸芯片对药物代谢酶多态性基因分型的优势。

Objective：To setup the protocol of multiplex PCR which suit the synchronizing hybridization and cutoff of signals discrimination. Methods： Optimisation of the primers and probes had been done; some key characteristics to PCR were adjusted and the final strategy was done. Preparing the oligoneueleid mieroarray, cutoff to determine wildtype and matant alleles was calculated. Results： It was recommended that ratios of wildtype allele to mutant signal which was higher than 4 or lower than 2.5 which be used as cutoffs in determination. When the ratio was in the range from 2.5 to 4, PCR or hybridization should be rerun. Conclusion：Multiplex PCR and synchronizing hybridization are helpful in epidermetology; the advantages of genechip is showed in CYP450s study by these techniques.