云南佤族、白族、傣族Y-SNP与Y-STR多态性研究与法医学意义
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摘要
[目的] Y染色体非重组区DNA多态呈父系遗传,在法医学混合斑分析、父系亲权鉴定及男性嫌疑人的案件排查中发挥重要作用。其单核苷酸多态具有种族、地理特异性,根据现场遗留男性个体生物学检材的特点来推测其种族地理来源可为侦查破案提供线索。本课题利用云南独特的少数民族遗传资源,选择了佤族、白族和傣族三个祖裔来源不同的民族进行具有东亚特异性的7个Y-SNP位点和Y filer试剂盒17个Y-STR基因座基因分型检测。获取了上述三个民族Y-SNP、STR两种遗传标记的基础数据分布特点,初步探讨了Y-STR与Y-SNP分型之间存在的关联性,及根据Y-SNP单倍群、Y-STR单倍型的特点来推测其种族来源的可行性,为更多民族群体利用Y染色体DNA多态研究及其在法医学领域中的应用提供一研究方法。
[方法]采集云南佤族(100例)、白族(146例)和傣族(158例)共404例无关男性健康个体的静脉血各5mL。采用酚-氯仿有机法提取核基因组DNA。用巢式PCR和PCR产物直接测序法进行7个Y-SNP分型检测；用Y filer试剂盒进行17个Y-STR基因座的复合扩增检测。应用Arlequin 3.5、Network4.5.1.6等软件进行数据统计分析。
[结果]所检测的7个Y-SNP,佤族在O3-M122、O2-M95、K*-M9、C-M130和F*-M89单倍群中有频率分布,无O1-M119和P*-M45分型；白族中7个Y-SNP单倍群均有分布；傣族除P*-M45外有其余6种单倍群分布。其中,O3-M122在佤族、白族和傣族中分别占56.0%、47.3%和31.0%；02-M95分别占15.0%、15.8%和39.9%；K*-M9分别占24.0%、13.0%和5.7%；C-M130分别占3.0%、10.2%和5.7%。17个Y-STR基因座构成的单倍型在佤族、白族和傣族中分别检出72种、115种和140种单倍型；三个民族之间无共有的单倍型：三个民族的Y-STR单倍型多样性分别为0.9875±0.0046、0.9957±0.0015和0.9981±0.0010。
佤族17个Y-STR基因座在各Y-SNP单倍群内均有高频率的等位基因分布,除DYS389Ⅱ和DYS19外,其余15个基因座的等位基因频率分布在03-M122和02-M95之间明显不同；DYS389 I, DYS19、DYS393、YGATAH4和DYS438中高频率等位基因在03-M122、02-M95和K*-M9单倍群之间的分布不同；03-M122的特征单倍型是15-12-23-29-15-15-12,17-12-10-11-22-13-13(14/12)-15-9-19,02-M95的特征单倍型是16-13-25-29-16-15-15,19-14-11-12-20-14-11-14-10-18。白族各Y-SNP单倍群内的等位基因频率分布比较均匀。傣族DYS389 I、DYS389 II、DYS19、DYS439、DYS392、DYS437、DYS438和DYS448高频率等位基因分布在03-M122和03-M95间不同,DYS390、DYS393和DYS635的等位基因在03-M122、03-M95和O1-M119单倍群之间分布不同。
同一Y-SNP单倍群的Y-STR单倍型Network网络图显示,佤族在03-M122、02-M95和K*-M9三个单倍群的网络图中均具有非常接近的单倍型并形成starcluster形状；白族在03-M122图中除部分单倍型位于中央外,单倍型之间差异比较大,02-M95网络图中具有一个小、star cluster形状；傣族在03-M122网络图中大部分的单倍型个体聚在一起,具有star cluster形状,且和部分白族个体的很接近,而在02-M95网络图中则各单倍型较分散,具有高度的单倍型多样性和分支多样性。
[结论]本课题研究获取了云南佤族、白族和傣族三个少数民族Y-SNP和Y-STR的群体遗传学基础数据,初步掌握了三个民族的父系遗传特点以及民族间的遗传差异。佤族的父系遗传结构单一,较好的保持了其父系遗传特征,其Y-STR等位基因频率分布与Y-SNP单倍群有相关性,在法医学案件检验中可利用其Y-STR特征单倍型及特异等位基因来推测其Y-SNP单倍群。而白族和傣族的父系遗传结构在多样性中存在着基因交流。
Objective:DNA polymorphism on non-recombined region of Y chromosome(NRY) play an important role in forensic casework such as mix stain analysis, paternity cases where the putative father or other relatives are lacking and cases of suspect lacking because of its patrilineal inheritance. The single nucleotide polymorphisms with geographic specificity provide the possibility to predict ethincgeographic origin of biological samples who left in the crime scene, which can help to provide scent in investigative phase of a criminal procedure. In this study, the subject based on the unique genetic resources of Yunan Province, the Wa, Bai and Dai which are descendants of three distinct ancient tribe were chosen, and 7 Y-SNPs with high polymorphism in Eastern Asia and 17 Y-STRs including in Y filer kit were typed. The data of Y-SNP and Y-STR of the distribution characteristics in three ethnic groups were obtained, the relatedness of alleles distribution at Y-STR locus and its Y-SNP haplogroups background and the feasibility of ethnic origin predicted from Y-STR haplotype were primary discussion. All these may be useful for further study of Y chromosome DNA polymorphism in other ethnic groups and application on forensic science.
Methods:Blood samples were collected from the Wa(n=100), Bai(n=146) and Dai(n=158), a total of 404 unrelated males were involved in this study. The genome DNA of 404 individuals were extracted with the phenol-chloroform method. All samples were typed using Nested amplified PCR and direct Sequencing methods with Y-SNP markers; 17 Y-STRs were typed using the AmpF/STR Yfiler system (Applied Biosystems). Data statistical analysis was used the software package Arlequin 3.5 and the program Network 4.1.5.6.
Results:Of the seven Y-SNP haplogroups, O3-M122, O2-M95, K*-M9, C-M130 and F*-M89 were found in the Wa while O1-M119 and P*-M45 were absent; All seven Y-SNP haplogroups were observed in the Bai and six were found in Dai expect P*-M45 haplogroup. The frequency distribution of O3-M122 in the Wa, Bai and Dai was occupied in 56.0%,47.3%and 31.0%respectively; O2-M95 was occupied in 15.0%,15.8%and 39.9%respectively; K*-M9 was occupied in 24.0%,13.0%and 5.7%and C-M130 was occupied in 3.0%,10.2%and 5.7%, respectively.72,115 and 140 different 17-loci haplotypes were found in Wa, Bai and Dai samples, respectively; No haplotype was shared among the three groups. Wa appeared in lowest haplotype diversity (0.9875±0.0046) compared to Bai (0.9957±0.0015) and Dai (0.9981±0.0010) groups.
In the Wa, high frequency distribution of alleles at 17 Y-STR loci was observed and showing striking difference between O3-M122 and O2-M95 haplogroups expect at DYS389Ⅱand DYS19; This difference was also found among O3-M122, O2-M95 and K*-M9 at DYS389 I, DYS19, DYS393, YGATAH4 and DYS438. Haplotype 15-12-23-29-15-15-12,17-12-10-11-22-13-13(14/12)-15-9-19 characterised in O3-M122 was sharply differ from haplotype 16-13-25-29-16-15-15,19-14-11-12-20-14-11-14-10-18 characterised in O2-M95. Alleles at most loci of the Bai showed moderate frequencies. In the Dai, the difference of high frequency distribution of alleles was observed between O3-M122 and O2-M95 at DYS389Ⅰ, DYS389Ⅱ, DYS19, DYS439, DYS392, DYS437, DYS438 and DYS448, and also observed among O3-M122, O3-M95 and O1-M119 at DYS390, DYS393 and DYS635.
In the median jioning network which constructed using Y-STR haplotypes under the same Y-SNP haplogroup. The Wa presented nearly identical haplotypes and displayed star-like features in network of O3-M122, O2-M95 and K*-M9; Bai showed relatively high diversity of haplotype in the O3-M122 network expect partly haplotypes which situating at center of the network, and a small star-like features was also observed in the network of O2-M95; The Dai displayed star-like features and closely distributed with some of Bai samples in the O3-M122 network, and showed high haplotype diversity and relatively scattered in the O2-M95 network.
Conclusions:In this study, the genetic data of Y-SNP and Y-STR in three ethnic groups were obtained, and the genetic charatistics of ethnic groups as well as explanation of the observed genetic differences amomg them were comparatively understood. Wa characterised in isolation ethnic and preserves a high level of patrilineal genetic diversity, and the allelic distribution at STR locus is strongly depend on its SNP haplogroups background which may have potential implications to predicte its Y-SNP haplogroup in forensic casework. In others, Dai and Bai revealed more intercommunion.
[方法]采集云南佤族(100例)、白族(146例)和傣族(158例)共404例无关男性健康个体的静脉血各5mL。采用酚-氯仿有机法提取核基因组DNA。用巢式PCR和PCR产物直接测序法进行7个Y-SNP分型检测；用Y filer试剂盒进行17个Y-STR基因座的复合扩增检测。应用Arlequin 3.5、Network4.5.1.6等软件进行数据统计分析。
[结果]所检测的7个Y-SNP,佤族在O3-M122、O2-M95、K*-M9、C-M130和F*-M89单倍群中有频率分布,无O1-M119和P*-M45分型；白族中7个Y-SNP单倍群均有分布；傣族除P*-M45外有其余6种单倍群分布。其中,O3-M122在佤族、白族和傣族中分别占56.0%、47.3%和31.0%；02-M95分别占15.0%、15.8%和39.9%；K*-M9分别占24.0%、13.0%和5.7%；C-M130分别占3.0%、10.2%和5.7%。17个Y-STR基因座构成的单倍型在佤族、白族和傣族中分别检出72种、115种和140种单倍型；三个民族之间无共有的单倍型：三个民族的Y-STR单倍型多样性分别为0.9875±0.0046、0.9957±0.0015和0.9981±0.0010。
佤族17个Y-STR基因座在各Y-SNP单倍群内均有高频率的等位基因分布,除DYS389Ⅱ和DYS19外,其余15个基因座的等位基因频率分布在03-M122和02-M95之间明显不同；DYS389 I, DYS19、DYS393、YGATAH4和DYS438中高频率等位基因在03-M122、02-M95和K*-M9单倍群之间的分布不同；03-M122的特征单倍型是15-12-23-29-15-15-12,17-12-10-11-22-13-13(14/12)-15-9-19,02-M95的特征单倍型是16-13-25-29-16-15-15,19-14-11-12-20-14-11-14-10-18。白族各Y-SNP单倍群内的等位基因频率分布比较均匀。傣族DYS389 I、DYS389 II、DYS19、DYS439、DYS392、DYS437、DYS438和DYS448高频率等位基因分布在03-M122和03-M95间不同,DYS390、DYS393和DYS635的等位基因在03-M122、03-M95和O1-M119单倍群之间分布不同。
同一Y-SNP单倍群的Y-STR单倍型Network网络图显示,佤族在03-M122、02-M95和K*-M9三个单倍群的网络图中均具有非常接近的单倍型并形成starcluster形状；白族在03-M122图中除部分单倍型位于中央外,单倍型之间差异比较大,02-M95网络图中具有一个小、star cluster形状；傣族在03-M122网络图中大部分的单倍型个体聚在一起,具有star cluster形状,且和部分白族个体的很接近,而在02-M95网络图中则各单倍型较分散,具有高度的单倍型多样性和分支多样性。
[结论]本课题研究获取了云南佤族、白族和傣族三个少数民族Y-SNP和Y-STR的群体遗传学基础数据,初步掌握了三个民族的父系遗传特点以及民族间的遗传差异。佤族的父系遗传结构单一,较好的保持了其父系遗传特征,其Y-STR等位基因频率分布与Y-SNP单倍群有相关性,在法医学案件检验中可利用其Y-STR特征单倍型及特异等位基因来推测其Y-SNP单倍群。而白族和傣族的父系遗传结构在多样性中存在着基因交流。
Objective:DNA polymorphism on non-recombined region of Y chromosome(NRY) play an important role in forensic casework such as mix stain analysis, paternity cases where the putative father or other relatives are lacking and cases of suspect lacking because of its patrilineal inheritance. The single nucleotide polymorphisms with geographic specificity provide the possibility to predict ethincgeographic origin of biological samples who left in the crime scene, which can help to provide scent in investigative phase of a criminal procedure. In this study, the subject based on the unique genetic resources of Yunan Province, the Wa, Bai and Dai which are descendants of three distinct ancient tribe were chosen, and 7 Y-SNPs with high polymorphism in Eastern Asia and 17 Y-STRs including in Y filer kit were typed. The data of Y-SNP and Y-STR of the distribution characteristics in three ethnic groups were obtained, the relatedness of alleles distribution at Y-STR locus and its Y-SNP haplogroups background and the feasibility of ethnic origin predicted from Y-STR haplotype were primary discussion. All these may be useful for further study of Y chromosome DNA polymorphism in other ethnic groups and application on forensic science.
Methods:Blood samples were collected from the Wa(n=100), Bai(n=146) and Dai(n=158), a total of 404 unrelated males were involved in this study. The genome DNA of 404 individuals were extracted with the phenol-chloroform method. All samples were typed using Nested amplified PCR and direct Sequencing methods with Y-SNP markers; 17 Y-STRs were typed using the AmpF/STR Yfiler system (Applied Biosystems). Data statistical analysis was used the software package Arlequin 3.5 and the program Network 4.1.5.6.
Results:Of the seven Y-SNP haplogroups, O3-M122, O2-M95, K*-M9, C-M130 and F*-M89 were found in the Wa while O1-M119 and P*-M45 were absent; All seven Y-SNP haplogroups were observed in the Bai and six were found in Dai expect P*-M45 haplogroup. The frequency distribution of O3-M122 in the Wa, Bai and Dai was occupied in 56.0%,47.3%and 31.0%respectively; O2-M95 was occupied in 15.0%,15.8%and 39.9%respectively; K*-M9 was occupied in 24.0%,13.0%and 5.7%and C-M130 was occupied in 3.0%,10.2%and 5.7%, respectively.72,115 and 140 different 17-loci haplotypes were found in Wa, Bai and Dai samples, respectively; No haplotype was shared among the three groups. Wa appeared in lowest haplotype diversity (0.9875±0.0046) compared to Bai (0.9957±0.0015) and Dai (0.9981±0.0010) groups.
In the Wa, high frequency distribution of alleles at 17 Y-STR loci was observed and showing striking difference between O3-M122 and O2-M95 haplogroups expect at DYS389Ⅱand DYS19; This difference was also found among O3-M122, O2-M95 and K*-M9 at DYS389 I, DYS19, DYS393, YGATAH4 and DYS438. Haplotype 15-12-23-29-15-15-12,17-12-10-11-22-13-13(14/12)-15-9-19 characterised in O3-M122 was sharply differ from haplotype 16-13-25-29-16-15-15,19-14-11-12-20-14-11-14-10-18 characterised in O2-M95. Alleles at most loci of the Bai showed moderate frequencies. In the Dai, the difference of high frequency distribution of alleles was observed between O3-M122 and O2-M95 at DYS389Ⅰ, DYS389Ⅱ, DYS19, DYS439, DYS392, DYS437, DYS438 and DYS448, and also observed among O3-M122, O3-M95 and O1-M119 at DYS390, DYS393 and DYS635.
In the median jioning network which constructed using Y-STR haplotypes under the same Y-SNP haplogroup. The Wa presented nearly identical haplotypes and displayed star-like features in network of O3-M122, O2-M95 and K*-M9; Bai showed relatively high diversity of haplotype in the O3-M122 network expect partly haplotypes which situating at center of the network, and a small star-like features was also observed in the network of O2-M95; The Dai displayed star-like features and closely distributed with some of Bai samples in the O3-M122 network, and showed high haplotype diversity and relatively scattered in the O2-M95 network.
Conclusions:In this study, the genetic data of Y-SNP and Y-STR in three ethnic groups were obtained, and the genetic charatistics of ethnic groups as well as explanation of the observed genetic differences amomg them were comparatively understood. Wa characterised in isolation ethnic and preserves a high level of patrilineal genetic diversity, and the allelic distribution at STR locus is strongly depend on its SNP haplogroups background which may have potential implications to predicte its Y-SNP haplogroup in forensic casework. In others, Dai and Bai revealed more intercommunion.
引文
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