The Research On The Polymorphisms Of The Two Loci,D6S1043 And D12S391 In Liaoning Han Population And The Method Of Multiplexing PCR

IntroductionSTR (short tandem repeats) refers to a kind of repetitive sequence whose length of repeat unit is 2-6bp. The length polymorphism depends on the different copy numbers of repeat unit between individuals. STR is the most commonly genetic marker used in forensic science currently, relying on its many advantages such as easiness in amplifying, accurate typing, fitness for challenged or degraded material, richness in genetic information, capability of multiplexing PCR and joint detection.Confronted with the growing number of forensic cases that are more and more complicated,in order to make a timely, accurate and authoritative appraisement, the kit including certain STR loci have been put into forensic application. Automated DNA typing has become the mainstream of current forensic science techniques, which makes use of multiplexing PCR and joint detection on the ground of STR kit. Since November 1997, the FBI (Federal Bureau of Investigation) established 13 core STR loci that are CSF1PO, FGA, TPOX, THO1, vWA, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51 and D21S11, a series of commercial kits including certain ideal STR loci have been widely used in personal identification, parentage testing and other important fields of forensic science characterized by accuracy, rapidness, convenience and delicacy. Investigating the genetic polymorphisms of the STR loci in different populations has become a focus of forensic biological evidence at present.Sinofiler Kit was developed by American ABI Company latest according to the genetic characteristics of Chinese population. D6S1043 and D12S391 are the two STR loci specially recommended by Sinofiler Kit, which are distinguished from the ones in other kits. This study investigated and obtained the genetic polymorphism data of the two loci in Liaoning Han population. The result could provide a theoretical foundation for the forensic application of Sinofiler Kit and a basis data for population genetics as well.Materials and MethodsGenomic DNA extraction and detection.According to the sequence published by U.S. National Center for Biotechnology Information database, synthesize the amplification primers of D6S1043 and D12S391 loci. After single-locus amplification, the genetic polymorphisms of two loci were detected by the polyacrylamide gel electrophoresis (PAGE). Collect and mix with the amplified products containing different alleles, combined with sequencing results to obtain allelic ladder. With the Ladder for reference, DNA fragments are separated by the method of PAGE and silver staining to determine genotypes on the two loci of 200 unrelated individuals in Liaoning Han population. Calculate the relevant forensic data. Regulate the two key factors of multiplexing PCR—nnealing temperature and the ratio of primer contents. Establish the method of multiplexing PCR about the two loci and analyse the genetic relationship between the three triad family samples by this method.Results1. In the 200 individuals of this study, there are 49 kinds of genotypes composed of 14 alleles (ranging from 9 to 22) on D6S1043 locus and 43 kinds of genotypes composed of 12 alleles (ranging from 15 to 26) on D12S391 locus.2. The forensic parameters of D6S1043 and D12S391 in Liaoning Han population are as follows.The values are 0.84 and 0.87 on heterozygosity (H),0.9651 and 0.9477 on discrimination power (DP),0.7363 and 0.6875 on the probability of exclusion (PE), 0.826 and 0.8499 on polymorphism information content (PIC) respectively.3. Establish the method of the multiplexing PCR about D6S1043 and D12S391. The best conditions of two key factors (annealing temperature and the ratio of primer contents) are 60℃and 0.8:1 respectively in this study. Conclusion1. The two loci, D6S1043 and D12S391 in Liaoning Han population are highly polymorphic STR loci and valuable in forensic application.2. There are national and regional differences in the distribution on genetic polymorphisms of D6S1043 and D12S391 loci, which is valuable in human genetics.3. The multiplexing PCR products of D6S1043 and D12S391 loci are capable of typing accurately by the method of PAGE combined with silver staining.