两种国产磁珠试剂盒对常见PCR抑制物去除效果比较

Comparative study on the removal effect of forensic common PCR inhibitors using two kinds of domestic magnetic beads kits

目的比较国产博坤磁珠试剂盒和Wawasye磁珠试剂盒对法医学常见PCR抑制物的去除效果。方法选择6种常见PCR抑制物,胆汁酸盐、胶原质、腐殖酸、血红素、黑色素和尿素,分别将其与K562 DNA混合后,采用Identifiler试剂盒进行分型,以恰好抑制K562 DNA所有26个不同等位基因的抑制物浓度为标准添加浓度,在此基础上提高每种PCR抑制物的浓度,分别采用两种磁珠试剂盒对混有抑制物的标准品DNA进行纯化处理后,再进行PCR扩增和STR分型检测,统计STR基因座等位基因检出个数。结果分别混有1×、2×和4×标准添加浓度各种抑制物的K562 DNA经博坤磁珠试剂盒纯化后,均可检出所有STR基因座的全部等位基因;经Wawasye磁珠试剂盒纯化后,混有胆汁酸盐、胶原质、血红素和尿素的DNA均可检出所有STR基因座的全部等位基因,而混有腐殖酸和黑色素的DNA未检出任何等位基因。结论除Wawasye磁珠试剂盒无法有效去除腐殖酸和黑色素外,两种国产磁珠试剂盒对大多数PCR抑制物均具有良好去除效果。

Objective To compare the removal effect of forensic common PCR inhibitors using domestic Bo Kun and Wawasye magnetic bead kits. Methods Six kinds of common PCR inhibitors,bile salts,collagen,humic acid,hemoglobin,melanin and urea were selected,and added respectively to the K562 DNA with standard addition concentration which just inhibit all of 26 different alleles of the K562 DNA with the Amp FSTR Identifiler PCR Amplification Kit. On this basis,after increased the concentration of each inhibitor and purified with two kinds of domestic magnetic bead kits respectively,PCR amplification and STR genotyping were performed,and then count the allele number of K562 DNA detected. Results After the K562 DNA mixed with 1 ×,2 × and 4 × standard addition concentration of various inhibitors was purified using the Bokun magnetic bead kit,all of 26 alleles of the K562 DNA could be detected with the Amp FSTR Identifiler PCR Amplification Kit. After purified with the Wawasye magnetic beads kit,all of 26 alleles of the K562 DNA mixed with bile acid,collagen,hemoglobin and urea could also be detected,and no allele could be detected,however,for those mixed with humic acid and melanin. Conclusion Both of two kinds of domestic magnetic bead kits can effectively remove the most of common PCR inhibitors except for humic acid and melanin with the Wawasye magnetic beads kit.