|
Short tandem repeat (STR) is the second generation genetic marker after the restriction fragment length polymorphism, since the late1980s when it was developed and used, it has got constant development in detection methods and applications. Now as the currently most widely used genetic markers, it is widely used in genetic mapping, gene mapping, diagnosis of genetic diseases, population genetics and forensic identification etc. The human Y chromosome, one of the sex chromosome, normal men have that but women do not, in addition to proposed autosomal region (pseudoautosomal region), restructuring does not occur in the other part during the the genetic process. The human Y chromosome STR (short tandem repeat) refers to short tandem repeat fragments exist in non-recombinant region on the Y-chromosome, structural features of the sequence can be stably passed from father to son, it is unique to men with the paternal inheritance.1976Cooke H first reported the presence of the human Y-chromosome tandem repeat sequence, has opened up a new way for the study of human genetics and forensic medicine. Arnemann found microsatellite DNA repeat units (gata)n which is named for DYS19now on the Y chromosome in1986, and other Y chromosome STR continued to be discovered since the1990s. Since the technology of PCR-STR set up, many domestic and foreign scholars have entered this field, a growing number of Y-STR polymorphisms are constantly being discovered. Currently,30mostly used Y chromosome microsatellite markers are DYS19, DYS287, DYS385, DYS388, DYS389Ⅰ, DYS389Ⅱ, DYS390, DYS391, DYS392, DYS393, YAC Ⅲ, Y-GATA-H4, Y-GATA-A4, Y-GATA-A7.1, Y-GATA-A7.2, Y-GATA-A8, Y-GATA-C4, Y-GATA-A10, DYS434, DYS435, DYS436, DYS437, DYS438, DYS439, DYS442, DYS446, DYS458etc. Some information of Y chromosome short tandem repeat are available in the U.S. population haplotype database (YHRD)(http://www.Ystr.Org/usa/) and European populations YHRD (http://ystr.Charite.de) obtained.Y-STR loci polymorphism have unique applications in the paternity testing of paternal family, the male component of the mixed plaque detection, determination the source of different males from a mixture of analysis and the body of the unknown man, paternal migration history and reconstruction of the same paternal family direction. Through the research of different racial, ethnic and geographic population genetics,Scholars found that the distribution of Y-STR polymorphisms have obvious differences during different racial, ethnic, geographical and family. The reason was during genetic process, in addition to pseudoautosomal region, the rest does not exchange and reorganize with the X chromosome, change in the sequence of this area is only caused by mutations, these mutations constitute polymorphisms in the non-recombinant part of the human Y chromosome. Accurate estimate of the mutation rate of the Y-STR loci in the study of human evolution is the premise to trace the lineage origin. The mutation rate of the Y-STR loci also explain exactly necessary for paternity testing and forensic findings.There are several ways to get Y-STR mutation:deep rooting pedigrees, multi-generation pedigree studies, the evolution of groups as well as single sperm cell research. In1997, Heyer et al studied42male individuals over12generation pedigree, a total of257meiosis were observed, remove the clear, suspicious of an illegitimate events in213meiosis,9Y-STR loci found four mutations, the mutation rate was2.1*10-3/generation. Because they were using a multi-generation pedigree, there would be non-paternity mutations, so that you could not absolutely determine the differences were caused by mutations or caused by a non-paternal inheritance, so of the mutation rate of Y-STR by multiple generations on pedigree also had been questioned. In2000, Kayser observed4999pairs of father and son (ie4999meiosis) and found14mutations in15Y-STR loci. The average mutation rate of Y-STR loci was2.8*10-3/generation, locus-specific mutation rates ranged from0to8*10-3/generation. Therefore, they believed that the mutation rate and the f characteristics of Y-STR were the same with autosomal microsatellite, and microsatellite mutation mechanism has nothing to do with the reorganization. Since then, Kurihara’ respectively, studied the mutation rate of the Y-STR loci of the different regions, and found the similar mutation rates (about2*10-3/generation). The mutation rate from different researchers had little difference between pedigree of the different regions. And they agreed that during the Y-STR mutation, the repeats number increased more than it decreased; long fragment of allele was tending to mutate compared to the short segment of alleles; most of the Y-STR mutation were one repeat difference, which were agreed with one step model. Now genotyping father-son pairs is the most reliable and direct method, but it require a large enough sample size. Few domestic research on the Y-STR mutation have been found, higher mutation rate observed in som studies after the detection of the Y-filer, around5%o, the reason may be related to the small sample size.2009Miriam observed1764meiosis between DNA analysis confirmed father and son pairs by using17Y-STRs in Yfiler, and detected84mutations, average mutation rate was2.5%o on each locus,98.8%of them were on step mutations,1.2%were two-step mutations, there was no significant differences between repeat gains and repeat loss, and there were a positive correlation between the Y-STR mutation rate and paternal age. Literatures reported that majority of Y-STR mutations have correlation with number of repetitions observed in the in the core sequence, and maybe correlated with differences between locus or allele. There is some controversy in the relationship of the Y-STR mutation rate and paternal age:Kayser found that there was no clear correlation between the mutation of the Y-STR and paternal age, while Gusmao found that Y-STR mutation rate increased with the increasing paternal age.The sharing point of the mechanism of Y-STR mutation is that during the process of DNA replication, DNA polymerase slips and results in the number of repeat in the core sequence change. STR loci mutation manifested as a change in the number of repeat units, mainly divided into two types of the former single repeat mutation and multiple repeat mutations, the former accounts for90%of the mutation events. About STR mutation mode, Brinkmann thought that single step mutation (ie, the change of a single repeat unit) accounted for more than90%of the STR mutation model through his own observations and combined with previous research. He proposed a step-by-step mutation model (step-wise a mutation model). Most of Y-STR mutation are one step observed by many Literature while minority are two step mutations. The mutation tends to increase or decrease in the number of repeat units, different literature give different conclusions.AmpFISTRmY-FilerTM is a colored fluorescent multiplex amplification system recently introduced by Applied Biosystems, amplification of the17loci can be synchronized in a reaction tube (DYS456, DYS389I D YS390, DYS389Ⅱ, DYS458, DYS19, DYS385, DYS393, DYS391, DYS439, DYS635, DYS392, Y-GATA-H4, DYS437, DYS438, DYS448). The kit has been widely used in forensic science case and human genetics research. The kit is based on the background information of white population, there is less information about the17Y-STR genetic parameters of the Chinese Han population, therefore, access to the17Y-STR loci of different groups in our country, especially polymorphisms in the Han population, mutation rate and characteristics of mutation, have very important value for the research and development of paternal paternity testing and Y-STR kit, Y-STR-based case investigation and paternal query system establishment.Our research genotyped1000Guangdong Han unrelated individuals,1000father-son pairs which have determine relationship by DNA testing, more than50pedigrees with more than three generations by Y-Filer kit. We observed the mutation phenomenon during17Y-STR transfer in meiosis, all mutation samples were sequenced to determine the nature of mutations, Y-STR mutation frequency in the Chinese Han population and mutation regularity. We assessd the value of17Y-STR in forensic medicine and provides theoretical support for paternal trace, development of Y-STR kit, pedigrees investigation and establishment of the platform based on Y-STR database, and to provide data for forensic science, population genetics, human evolution research.A direct counting method is used to to calculate allele frequencies in Statistical analysis, the i-th frequency of the alleles of the locus with Pi=the i-th number of allele frequency/n (n is the quantity of samples for the group). Genetic diversity (gene diversity, GD) and haplotype diversity (haplotype diversity, HD) are calculated according to the formula h=n of (1-ΣPi2)/(n-1), n is the quantity of sample cases, Pi is allele frequency.Conclusion:No men shared the same haplotype of17Y-STR loci in the survey of1000the Guangdong Han Chinese men unrelated individuals. HD (haplotype discrimination) reached to1.0000. All locus genotype frequency were shown in Table1. There were two copies (a and b) in DYS385locus, the haplotype frequencies were shown in Table2. The GD (Gene diversity) of17Y-STR loci were between0.4285(DYS391) to.9654(DYS385a/b); DYS385a/b were found81genotypes, including five tri-allele patterns and one sample with four alleles (see Table2); the the rest locus were found in5to10genotypes. Our study also observed1000meiosis between father and son,46mutations were found in11loci (ie, DYS458, DYS439, DYS385a/b, DYS389II, DYS390, DYS635, DYS456, DYS392, DYS19, DYS389I), all mutations were confirmed by sequencing analysis. Average mutation rate of17loci was0.0027(95%CI,0.0020-0.0036). Among these46mutations,16mutations (i.e.34.78%) gained repeats and30mutations (i.e.65.22%) lost repeats, the ratio was1:1.87. Longer allele got higher mutation rate than the short and median alleles; Y-STR mutation rate of older father was almost2times higher than mutation rates for younger father. In a total50pedigrees of Su and Yang surname,30Su-surnamed families have10haplotypes;20Yang-surnamed pedigrees had14haplotypes. Su-Surnamed pedigrees were from five different paternal haplotype; Yang-surnamed pedigrees were from the two paternal haplotype, no shared haplotype are found in these two surname pedigrees.
|