一起死亡事故中难辨尸体的个体基因识别

目的对某地一起死亡事故中难辨认尸体进行个体基因识别。方法从遇难者尸体取深部肌肉和软骨组织少许。对部分组织进行常规病理检查。从尸体组织提取基因组DNA，采用荧光多重PCR技术，扩增尸体基因组DNA的16个短串联重复序列（STR）基因座。将遇难者STR基因型与父母的STR基因型进行比对，确定遇难者的家庭归属。结果常规病理检查示部分肌肉组织结构完全破坏，但软骨组织结构尚完整。肌肉组织基因组DNA的凝胶电泳呈3种类型：基本无降解、部分降解、完全降解。只有部分肌肉组织DNA可用于确定STR基因型，所有软骨的DNA样本均可用于STR-PCR。结论以STR基因型分析为基础的亲子鉴定技术是确定事故遇难者身份的有效方法。对腐烂尸体的STR分析而言，尸体软骨组织是首选。     

表皮松解性掌跖角化症家系基因突变研究

目的确定一个表皮松解性掌跖角化症家系的致病基因。方法收集该家系3例患者、3例表型正常个体和50名无亲缘关系的健康个体的外周血标本,抽提基因组DNA。选取角蛋白9(KRT9)基因邻近的3个STR多态性位点D17S1787、D17S579、D17S250进行连锁分析研究,并对KRT9基因所有外显子进行PCR扩增和Sanger测序分析。结果 3个STR位点均与患者表型共分离;患者的KRT9基因第1外显子均可见c.488 G>A(p.R163Q)杂合性突变,而家系中表型正常个体和50名正常对照个体均未检测到KRT9基因突变。结论 KRT9基因的c.488 G>A错义突变是导致该家系发生表皮松解性掌跖角化症的原因。     

常染色体显性遗传先天性板层白内障家系致病基因的排除性定位

目的 定位-个四代常染色体显性遗传先天性板层白内障家系的致病基因.方法 选取在北京同仁医院就诊的河北任丘先天性白内障家系,记录家系遗传史.该家系28例成员(12例患者,16例非患者)进入本研究,12例患者接受全身及眼部检查,以排除存在白内障以外的眼部及全身疾患,16例非患者仅接受眼部检查.28例研究对象均采集外周静脉血5ml,提取基因组DNA,选取在物理距离上与已知非综合征常染色体显性遗传性先天性白内障相关的18个致病基因紧密连锁的微卫星分子标记,基因组聚合酶链式反应(PCR)扩增后进行基因分型.以基因分型的结果为基础,利用等位基因共享分析和基因测序对已知候选基因进行排除定位.结果 该家系遗传特点符合常染色体显性遗传,临床表型为板层先天性白内障;与位于1、2、10、11、12、16、17、21、22染色体上的15个致病基因附近的微卫星位点均不存在等位基因共享,基因测序排除了微卫星杂合度较低(位于3、13,19号染色体)的基因座.结论 该家系存在新的致病基因,进一步确证了先天性白内障具有高度临床和遗传异质性.该家系致病位点的确定有待于进一步研究.     

两种全血基因组DNA提取方法的比较

目的比较两种不同的DNA提取方法提取人全血基因组DNA,对DNA提取率、纯度以及PCR扩增的效果影响.方法分别用传统的酚、氯抽提法和改良法两种不同的方法,提取人全血基因组DNA.结果两种方法提取的DNA纯度用光吸收比值为指标检测A280/260值为:1.45和1.82,提取DNA总量分别为187μg/ml和260 μg/ml.用0.8%琼脂糖凝胶电泳鉴定改良法制备的DNA效果较好,PCR扩增效果有差异.结论用改良法提取的DNA总量明显较高,提取效率高,PGR扩增的效果好,是全血基因组DNA提取中首选考虑的方法.     

慢性阻塞性肺疾病免疫学指标的变化及其与DNA短串联重复序列的关联

目的探讨慢性阻塞性肺疾病(COPD)患者血液免疫学指标与DNA短串联重复序列的关联性。方法选取COPD患者40例及健康对照组100例应用全自动生化分析仪测定血液IgA、IgG、IgM、C3、C4,并应用第二代DNA遗传标记-短串联重复序列(STR)对D8S1179、D21S11、D7S820、CSF1P0、D3S1358、D5S818、D13S317、D16S539、D2S1338、D19S433、VWA、D12S391、D18S51、D6S1043、FGA 15个STR基因位点的遗传多态性进行分析。结果 COPD组免疫学指标与对照组比较,C4、IgA明显升高(P<0.05);而IgM升高、C3和IgG降低(P>0.05)。其中IgM与D8S1179-12、D19S433-16.2位点明显相关(P<0.05),IgG与FGA21位点明显相关(P<0.05)。结论 COPD患者免疫学指标发生变化,推测IgM、IgG检测结果与基因位点有关联,可能由基因决定了免疫学指标的个体差异性。     

大鼠NR2B基因核心启动子区单核苷酸多态性研究

 目的:对大鼠NR2B基因核心启动子区进行单核苷酸多态性(single nucleotide polymorphism,SNP)筛查及分析.方法:提取16只健康SD大鼠海马组织基因组DNA,聚合酶链式反应(PCR)扩增NR2B基因核心启动子区序列和SNP库已报道的rs8169392所在区域序列,结合DNA测序方法进行SNP筛查和验证.结果:在大鼠NR2B核心启动子区705bp中,发现1个新的SNP位点,为-217位C/T多态;本研究中未筛查到NCBI-SNP库报道的rs8169392.结论:-217位C/T多态的发现为NR2B基因多态性数据库提供了新信息.     

不同居群白芍的DNA随机扩增多态性研究

目的分析6个不同居群白芍的遗传多样性,为白芍的种质鉴定及遗传多样性分析提供依据。方法运用随机扩增多态性DNA（RAPD）技术,对浙江磐安、四川中江、安徽亳州、上海崇明、江苏宿迁和山东荷泽居群白芍的基因组DNA进行随机扩增,利用NTsys2.10e软件计算遗传相似性,运用UPGMA法进行聚类分析并构建树状图。结果共筛选了70个随机引物,从中挑选出8条多态性强、重复性好的引物,共检测出215个位点,多态性位点137个,多态位点比率为63.7%,UPGMA聚类可以将不同来源的白芍很好地区分开。结论不同产地间的白芍存在丰富的遗传多样性,RAPD分子标记方法可以用来鉴定不同产地的白芍。     

不同能量质子物理作用过程致DNA损伤的对比研究

目的 基于蒙特卡罗程序包Geant4研究布拉格峰附近能量质子在物理作用过程阶段对DNA损伤的特点规律。方法 利用Geant4构建细胞核及DNA模型,并基于Geant4-DNA物理过程模拟布拉格峰附近7档不同能量的质子在细胞核中传输过程,记录各相互作用位点相关信息,随机选取16%作用位点作为质子及其次级电子与DNA相互作用的位点,并将可导致DNA链断裂的位点信息写入新的文件,之后通过基于密度的空间聚类算法（DBSCAN）处理该文件,进而分析计算不同能量质子致DNA损伤的差异。结果 能量介于0.6～20.0 MeV之间的质子穿过细胞核时,随着能量的降低,单个质子致DNA损伤位点数由49.86增至549.88;损伤集簇数由2.92增至82.46;各尺寸集簇数增加显著,尺寸≥5的集簇增加329倍以上;平均集簇尺寸虽也有增加,但并不明显;简单单链断裂、复杂单链断裂、简单双链断裂和复杂双链断裂分别增加约7、25、23和63倍;单链断裂所占比例由96.69%减少到89.37%,双链断裂则由3.31%增加至10.63%。结论 质子能量越低,通过物理作用过程使DNA产生的损伤越复杂,DNA的修复越困难。     

青海蒙族群体8个STR位点遗传多态性研究

目的：研究青海蒙族人群第21号染色体D21S1432、D21S1435、D21S1270、D21S1440、D21S1446、GATA24H09、ATA42C09、GATA129D11等8个STR位点的遗传多态性。方法：运用PCR扩增、6%变性聚丙烯酰胺凝胶电泳结合银染技术对30位无关个体蒙族人群进行多态性研究。结果：8个位点分别检测出6、5、7、5、6、5、5、5个等位基因片段,共140个基因型,频率分布在0.025 09~0.046 43之间,多态性分布符合Hardy-weinberg平衡定律。8个STR位点多态信息量（PolymorpHism information content,PIC）分别为0.699 4、0.703 9、0.759 2、0.638 0、0.672 0、0.685 0、0.674 0、0.691 9,累积多态信息量为0.688 9。期望杂合度（heterozygosity,HET）分别为0.740 2、0.748 3、0.790 4、0.682 2、0.741 5、0.7271、0.742 5、0.725 5,累积杂合度为0.758 9。累积个体识别率（discrimination power,DP）为0.989 6,累积排除率（probabilities of paternity exclusion,PE）为0.999 6。结论：青海蒙族8个位点STR基因座的多态性数据显示其基因分布特征具有特异性。     

双胎胎儿畸形的磁共振诊断

目的:评价MRI对胎儿双胎和双胎畸形的诊断价值.方法:回顾性分析经引产或手术证实的5例双胎产前磁共振成像,并将产前MRI、产前超声(US)结果与引产后或出生后随访结果对照.结果:5例双胎中,产前MRI诊断连体双胎1例,双胎四肢短肢畸形1例,双胎小肠狭窄1例,双胎一胎无心无脑畸形1例,双胎一胎无脑畸形1例.结论:MRI能较好的诊断胎儿双胎及双胎畸形,由于其大视野,较好的软组织对比度,且不受羊水量多少、孕妇体型、胎儿位置、胎儿颅骨及母体骨盆骨骼等因素影响.     

Assessment of application value of 19 autosomal short tandem repeat loci of GoldenEyeTM 20A kit in forensic paternity testing

This study was carried out to assess the application value of 19 autosomal short tandem repeat (STR) loci of GoldenEye^TM 20A kit, in which 13 combined DNA index system core STR loci and PentaE, PentaD, D2S1338, D19S433, D12S391, and D6S1043 of six STR loci could be used in forensic paternity testing in Chinese population. We amplified the genomic DNA from blood samples on FTA paper of 289 paternity testing cases by using the GoldenEye^TM 20A kit. The amplified products were detected by capillary electrophoresis, and then the genotypes of 20 genetic markers including 19 STR loci as well as Amelogenin for sex determination were analyzed by GeneMapper v3.2 and GeneMarker HID Software. The results of genotypes were compared to the three commonly used commercial kits including AmpF?STR Identifiler^TM, PowerPlex^TM16, and AmpF?STR Sinofiler^TM kits. Compared to the three other common commercial kits, the GoldenEye^TM 20A kit had higher value of combined paternity index in certainty of paternity or non-exclusion paternity cases, and more numbers of STR loci were excluded in exclusionary paternity cases. Our data in this study showed that the GoldenEye^TM 20A kit has a higher application value in forensic paternity testing and will be of help for kinship analysis.     

Parentage testing with 14 STR loci and population data for 5 STRs in the Slovenian population

In order to apply a set of 14 short tandem repeat (STR) loci in parentage testing, we performed a population genetic study on a sample of 260 unrelated people from the Slovenian population. Genotypes for the 14 STRs were determined using three multiplex polymerase chain reactions (PCR) and automated fluorescent detection. The allele frequencies of the STR loci D5S818, D13S317, D7S820, D8S1179 and D18S51 showed no deviation from the Hardy-Weinberg equilibrium and agreed well with other Caucasian populations. We resolved a series of 181 parentage disputes of which 29 were exclusions. In all cases, evidence for exclusion was obtained by at least 4 informative STRs out of the 14 loci analysed. The 14 loci combined comprise a highly discriminating test suitable for paternity and identity testing in the Slovenian population, with an average estimated mutation rate of 1.2x10(-3), a combined calculated power of exclusion of 99.99974% and paternity index (PI) value of >10(6) in 72% of the inclusion cases and >10(5) in 91% of the inclusion cases.     

The identification of war victims by reverse paternity is associated with significant risks of false inclusion

Since February 2001 the process of DNA identification of war victims in Croatia relies on the database of over 3,000 9-locus (D3S1358, vWA, FGA, TH01, TPOX, CSF1PO, D5S818, D13S317 and D7S820) STR genotypes of relatives of missing persons. Instead of a targeted approach to DNA typing, the genotype of each skeletal remains analysed is compared to all genotypes in the database to identify potential parents and children. Although this approach has significantly increased the pace of identification by DNA typing, non-targeted matching in a database containing several thousand genotypes considerably decreases the significance of inclusion, especially when identification is based on reverse paternity analysis. To support this statistical prediction we present 3 cases of 10 STR loci matches and 1 case of 11 STR loci matches between a child, child's mother and skeletal remains that did not originate from a father of that child.     

Pseudo-exclusion from paternity due to maternal uniparental disomy 16

The investigation of a case of disputed paternity revealed indirect exclusion of the alleged father in the haptoglobin system and in the DNA single-locus system D16S309/Hinf I (MS205). The paternity index for the non-exclusion systems was > 10⁶. Since both exclusion systems (HP and MS205) are located on chromosome 16, we investigated 10 microsatellite loci covering this chromosome with 10–20 cM resolution. Analysis of the child’s chromosome showed only alleles of maternal origin and lack of inheritance of paternal alleles for five informative loci. The markers close to the centromere of chromosome 16 were heterozygous, whereas distal loci were either heterozygous or homozygous for maternal alleles. This is consistent with a maternal meiosis I nondisjunction of chromosome 16 leading to maternal uniparental heterodisomy. This case emphasizes that the opinion of non-paternity should be based on the absence of paternal alleles at genetic systems located on at least two different chromosomes. Received: 23 December 1997 / Received in revised form: 9 February 1998     

Genetic data on 10 autosomal STR loci in the Bangladeshi population

Allele frequencies of 10 autosomal short tandem repeat (STR) loci, D3S1358, vWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01 and FGA were determined in 211 unrelated Bangladeshi individual using AmpFLSTR SGM Plus PCR Amplification Kit. Statistical parameters of forensic importance, the power of discrimination (PD), observed and expected heterozygosity values (H), polymorphism information content (PIC), probability of match (PM), power of exclusion (PE) and typical paternity index (TPI) were calculated for the loci. These parameters indicated the usefulness of the loci in paternity testing and personal identification in the Bangladeshi population.     

Loss of heterozygosity detected at three short tandem repeat locus commonly used for human DNA identification in a case of paternity testing

Short tandem repeat (STR) is widely used for DNA profiling in forensic sciences for its stable inheritance. Genomic variations in STR loci may affect the results of the genotyping. In this study, using STR profiling and genome-wide chromosomal microarray assay, we detected the incidence of uniparental disomy or copy-neutral loss of heterozygosity (LOH) in a case of a parental testing, which altered the genotype of three commonly used STR markers including D2S1338, D2S441 and D2S1776. To the best of our knowledge, this is the first time found that LOH affect the genotyping of STR markers commonly used for paternity testing. Our findings demonstrated that the incidence of LOH in the genome may dramatically alter the results of DNA identification, and suggested that genomic structure variation need to be taking into consideration in the DNA identification using STR markers.     

Population data and mutation rates of 19 STR loci in seven provinces from China based on Goldeneye™ DNA ID System 20A

Short tandem repeat (STR) analysis is a primary tool in forensic casework. Population data and mutation rates of STRs are very important for paternity testing and forensic genetics. However, the population data and mutation rates of STRs in Han nationality based on large samples have still not been fully described in China. In this study, the allelic frequencies, forensic parameters, and mutation rate of 19 STR loci (D19S433, D5S818, D21S11, D18S51, D6S1043, D3S1358, D13S317, D7S820, D16S539, CSFIPO, PentaD, vWA, D8S1179, TPOX, Penta E, TH01, D12S391, D2S1338, and FGA) based on the Goldeneye™ DNA ID System 20A in Southern China Han nationality among seven provinces were investigated. Furthermore, population stratification of Southern China Han nationality among seven provinces was established. The multidimensional scaling (MDS) plot based on genetic distances (Fst) showed that the studied populations can be clustered into two major groups. However, relationships among populations were weak (Fst < 0.0043). A total of 376 cases of mutation were detected from the 19 selected loci in 15,396 meioses. The average mutation rate for the 19 loci was estimated to be 1.3 × 10⁻³ per meiosis. The mutation was mainly single step; the paternal mutation rate was higher than the maternal; and paternal mutation rate increases with paternal age.

     

Paternity testing after pregnancy termination using laser microdissection of chorionic villi

We report an unusual case of paternity testing from residues of chorionic villi 5 weeks after pregnancy termination. The autopsy of a 32-year-old female homicide victim revealed the presence of intact chorionic villi at the former placenta implantation site. Fetal cells were selectively isolated by laser-induced microdissection of the remaining villi to avoid contamination with maternal DNA. Simultaneous amplification of 12 STR loci in 2 PCR reactions resulted in a combined probability of paternity of 99.94%. This case demonstrates that laser-assisted microdissection and multiplex STR typing provide tools for paternity testing performed on endometrial mucosa long after the product of conception was removed by therapeutic abortion. Received: 2 May 2000 / Accepted: 7 November 2000     

Fourteen non-CODIS autosomal short tandem repeat loci multiplex data from Taiwanese

Interest in the development of polymorphic short tandem repeat (STR) markers unlinked to the CODIS loci is growing among forensic practitioners. We developed a multiplex system in which14 autosomal STR (D3S1744, D4S2366, D8S1110, D12S1090, D13S765, D14S608, Penta E, D17S1294, D18S536, D18S1270, D20S470, D21S1437, Penta D, and D22S683) could be amplified in one single polymerase chain reaction. DNA samples from 572 unrelated Taiwanese Han subjects were analyzed using this 14 STR multiplex system. Thirty parent–child pairs of parentage testing cases with a combined paternity index (CPI) below 1,000 and 32 parent–child pairs with single-step mutations found in AmpFℓSTR Identifiler loci were also recruited for validation of the newly developed system. DNA sequencing was performed for novel STRs and novel alleles found in these subjects. The distributions of allelic frequencies for these autosomal STRs and sequence data, allele nomenclature for the STRs, and forensic parameters are presented. The discrimination power in our multiplex loci ranged from 0.6858 (D18S536) to 0.9168 (Penta E), with a combined discrimination power of 0.999999999. It provides additional power to distinguish the possible single-step mutations in parent–child pairs and improves the ability to prove parentage by increasing the CPI. The combined power of exclusion of these 14 loci in Taiwanese Han in this study was 0.9999995913. In conclusion, this 14-autosomal STRs multiplex system provides highly informative STR data and appears useful in forensic casework and parentage testing.     

The new guidelines for paternity analysis in Germany—how many STR loci are necessary when investigating duo cases?

The requirements in the new German guidelines for paternity analysis have not only changed according to the so-called Gendiagnostikgesetz, the new German law regulating human genetic as well as paternity analyses, but also regarding the minimal number of short tandem repeats (STRs) which should be investigated (15 STRs) and the minimal required average exclusion chance (99.999 %). Even in paternity analyses involving only two people (e.g., father and child or mother and child), this exclusion chance is mandatory. A retrospective analysis of 330 father–child cases from our routine investigations showed in 142 cases (43 %) an individual exclusion chance below 99.999 % when using 15 STRs as required, in our routine work provided by the Powerplex® 16 kit which is reported to have an average exclusion chance of 99.988 %. Therefore, these same 330 father–child pairs were additionally analysed using the Powerplex® 21 kit and 120 of these duos were additionally analysed using the Powerplex® ESX17 kit enabling the analysis of 20 or 16 loci respectively. Now, an individual exclusion chance of more than 99.999 % could be achieved in 95.5 % (Powerplex® 21; calculation without the results of D6S1043), 98.8 % (Powerplex® 21; calculation with the results of D6S1043, using allele frequencies established in this study for a German and a West African population) and 98.3 % (Powerplex® ESX17). These data clearly demonstrate that in duo cases, more than the required 15 STR loci have to be investigated to obtain sufficient results.