联苯胺试验对血痕DNA检验的影响

摘要

Objective To explore the effect of benzidine test and related reagents on DNA analysis of bloodstain. Methods A total of 970 bloodstain filter paper samples with 1μL venous blood were collected, and 10 of them acted as control samples. After benzidine test and related reagent processing, DNA of 960 samples was extracted by Chelex-100 and silica bead methods and then multiplex amplified by AmpF(e)STRTM IdentifilerTM Plus PCR kits. The results of STR typing were compared between different groups. Results DNA were extracted immediately after benzidine test. Totally STR loci (3.80±1.34) were detected by silica bead method, while no STR loci were obtained by Chelex-100 method. Thirteen sam-ples (21.7％) with whole STR typing results were obtained by drying after benzidine test, and the STR locus number (12.90±1.49) which obtained by silica bead method was much higher than by Chelex-100 method (4.70±1.96) (P<0.05). When DNA was extracted immediately after the addition of glacial acetic acid, the STR locus number was (9.40±2.09) by silica bead method, but no STR typing result was obtained by Chelex-100 method. All 15 STR loci could be obtained by only adding glacial acetic acid after drying and only adding tetramethylbenzidine alcoholization liquid or 3％ hydrogen peroxide liquid. Conclusion Benzidine test has significant influence on DNA analysis of bloodstain. The Chelex-100 method is not suitable for the DNA extraction of bloodstain after benzidine test. Drying after benzidine test and silica bead methods can effectively enhance the STR locus number of bloodstain.%目的 探讨联苯胺试验及相关试剂对血痕DNA检验的影响.方法 制作含1μL静脉血的滤纸血痕970份,其中10份为对照样本,960份经联苯胺试验及相关试剂分别处理后,采用Chelex-100法和硅珠法提取DNA,AmpF詛STRTM IdentifilerTM Plus PCR扩增试剂盒进行复合扩增,对比各组STR分型结果.结果 联苯胺试验后立即提取DNA,硅珠法的STR基因座检出数为(3.80±1.34)个,而Chelex-100法均未获得STR分型结果;联苯胺试验后干燥处理,硅珠法有13例(21.7％)获得全部STR基因座分型结果,且STR基因座检出数[(12.90±1.49)个]远高于Chelex-100法[(4.70±1.96)个](P<0.05);加入冰醋酸后立即提取DNA,硅珠法的STR基因座检出数为(9.40±2.09)个,而Chelex-100法均未获得STR分型结果;只加冰醋酸后干燥处理以及只加四甲基联苯胺乙醇饱和溶液或3％过氧化氢溶液,两种方法均获得完整15个STR基因座分型结果.结论 联苯胺试验对血痕的后续DNA检验有很大的影响,Chelex-100法不适合联苯胺试验后血痕的DNA检验,联苯胺试验后干燥处理及采用硅珠纯化的方法可有效提升联苯胺试验后血痕的STR基因座检出数.