Maxwell 16裂解纯化法提取陈旧精子DNA的应用

目的利用Maxwell 16裂解纯化法从保存8年以上陈旧精斑检材中获取精子DNA。方法 8份陈旧精斑检材采用Maxwell 16裂解纯化法提取精子DNA,并采用Powerplex○R21试剂盒进行复合扩增,产物用AB3130型遗传分析仪检测,结果与常规方法进行对比。结果成功获得8份陈旧精斑检材精子STR分型。结论差异裂解配合Maxwell 16裂解在陈旧精斑检材精子DNA检验中效果明显。     

常染色体STR及线粒体测序用于陈旧性骨骼DNA一例

陈旧性骨骼是法医学实践中一种检验难度比较大的生物检材,常常由于DNA的降解造成检验的失败。本文通过优化陈旧性骨骼的DNA提取条件,对4例埋葬38年的尸骨进行了STR分型和线粒体测序检验．为当事人成功解决了已故亲人尸骨的认定。现报道如下。     

超滤管在陈旧生物检材中的应用

目的探讨millipore超滤管滤过方法对陈旧生物检材DNA分型检验的应用价值。方法将23份陈旧血样分别剪取同样合适大小的血片3组,标为A、B、C组,磁珠法提取DNA,分别用80μL、80μL、20μL洗脱液洗脱得到模板DNA,其中A、C组模板直接扩增,B组用millipore超滤管滤过浓缩后扩增。PCR产物用AB3130x L基因分析仪检测,Gene Mapper ID V3.2软件进行自动分型。所得实验数据用SPSS软件分析处理。结果 A组没有1例样本扩出全部STR基因座,B组有18例样本扩出全部STR基因座,C组有11个样本扩出全部STR基因座。结论应用millipore超滤管滤过方法可以明显提高陈旧生物检材的DNA分型成功率。     

衣物类生物检材DNA检验的研究

目的建立对衣物类生物检材STR检验的有效自动提取纯化方法。方法对100例衣物类生物检材用EZ1DNA Investigator试剂盒、EZ1Advanced XL工作站的Large-Volume程序(大体积法)进行提取纯化,AmpFLSTR~Identifiler~Plus荧光STR复合扩增,3500x L型遗传分析仪分析结果。结果 69份检材中检出了单一DNA分型结果,9份检出了混合基因型,22份未检出基因型。结论该方法对衣物类生物检材DNA提取纯化后进行荧光STR检测,可应用于法庭科学实践。     

常染色体及 Y染色体 STR分型技术在检案中的应用

荧光标记 STR分型技术所需检材量少，对陈旧、腐败检材适用性强，结果准确、灵敏度高，而 Y染色体 STR分型技术为男性特有，且 Y染色体 STR基因座不与其他染色体发生交换，在一些特殊案件的侦破中，联合应用常染色体及 Y染色体 STR分型技术往往可以起到关键性的作用。本文结合一典型系列案例进行分析，对案件检材提取、检验结果及应用价值等进行分析讨论，旨在提高 DNA检验技术在同类案件中的应用价值。 1 案 情 　　某年 6月至 11月，某市发生女青年搭乘摩托车到郊区偏僻地点被车手抢劫、强奸的系列案件共 21宗，送检的被害人内裤…     

荧光标记短片段STR复合扩增系统的法医学应用研究

目的解决严重降解(低于300bp)DNA检验问题,提高降解DNA的检出率。方法重新设计引物,减小扩增产物片段长度,用荧光标记短片段STR复合扩增体系进行DNA检验,扩增结果与用Identifiler试剂盒扩增出的结果进行比较。结果用荧光标记短片段STR复合扩增系统对实际案件中的高度降解DNA检材进行检验,可以获得满意分型。结论该系统可用于严重降解DNA检材的检验工作。     

两种提取骨骼、牙齿DNA的方法

目的建立提取骨骼、牙齿DNA的新方法。方法收集380例骨骼、牙齿检材,其中347份为常规骨骼、牙齿(常规组),33份为陈旧骨骼、牙齿(疑难组)。常规组检材以Handy-Eco仪器、疑难组则用冷冻研磨仪研磨成粉,以Kingfisher自动化系统提取DNA,用Identifiler Plus进行STR分型检测。结果用HandyEco Kingfisher法(H-K法)成功提取345例常规骨骼、牙齿检材的DNA,成功率99.42%;用Freeze-mill Kingfisher法(FM-K法)成功获取32例疑难骨骼、牙齿检材的DNA,成功率96.97%。结论采用H-K法和FM-K法对骨骼和牙齿DNA的检验成功率较高,可选择应用于工作实践中。     

联合应用chelex100和磁珠法检测霉变皮上衣微量DNA1例

微量疑难检材DNA的提取是目前检案的难点之一.笔者在办案中,遇到1份微量、陈旧、霉变的检材;经联合应用chelex100法和磁珠法(DNA IQIM)成功从检材中提取并获得了足量、高纯度的模板DNA,采用PCR扩增和荧光检测获得理想的STR分型结果,为破案提供可靠有力的科学依据.现报道如下.     

接触性检材502胶熏显后手印脱落细胞的DNA提取

目的建立接触性检材中手印定位、检材转移、DNA提取纯化的方法。方法 66例接触性检材经502胶熏显,手印接触部位明确后,分别采用普通擦拭法及丙酮擦拭法进行检材上手印脱落细胞的转移,并使用超微量磁珠法试剂盒提取纯化手印脱落细胞DNA,扩增后分析结果。结果用丙酮擦拭法得到的33例检材有30份获得了较好的STR分型结果,峰值范围为1 000~4 000 RFU,峰型均匀;而普通擦拭法很难获得理想的STR分型。结论通过丙酮擦拭法转移502胶熏显的手印脱落细胞,STR分型效果更优良,可应用于法庭科学实践。     

强奸案中的DNA证据

强奸案中DNA证据有哪些作用1．确定案犯确定案犯是DNA证据首要的也是最重要的作用。目前用于法医DNA分析的STR基因座在人类基因组中分布广泛,适用于陈旧降解及微量生物检材的分析。以目前常规用于DNA检测的16个STIR基因座复合扩增试剂盒而言,除同卵双生子之外,没有两个人的DNA分型结果一致。所以,通过对现场犯罪分子遗留的精斑、唾液、血迹等生物物证的检测分析并与嫌疑人进行比对,就可以直接认定。STR基因座判型准     

High‐Resolution Melting (HRM) of Hypervariable Mitochondrial DNA Regions for Forensic Science

Forensic strategies commonly are proceeding by analysis of short tandem repeats (STRs); however, new additional strategies have been proposed for forensic science. Thus, this article standardized the high‐resolution melting (HRM) of DNA for forensic analyzes. For HRM, mitochondrial DNA (mtDNA) from eight individuals were extracted from mucosa swabs by DNAzol reagent, samples were amplified by PCR and submitted to HRM analysis to identify differences in hypervariable (HV) regions I and II. To confirm HRM, all PCR products were DNA sequencing. The data suggest that is possible discriminate DNA from different samples by HRM curves. Also, uncommon dual‐dissociation was identified in a single PCR product, increasing HRM analyzes by evaluation of melting peaks. Thus, HRM is accurate and useful to screening small differences in HVI and HVII regions from mtDNA and increase the efficiency of laboratory routines based on forensic genetics.     

Application of mtDNA SNP analysis in forensic casework

In this study six forensic cases are presented where the routine analysis of samples for short tandem repeats (STRs) failed. The sequencing of the mitochondrial hypervariable region I (HVR I) also failed. Nevertheless, it was possible to analyse the samples with mitochondrial DNA (mtDNA) single nucleotide polymorphisms (SNPs) via SNaPshot technique. The age of the analysed samples ranged from 2 months to 1400 years. Saliva-, blood-, sperm-, hair-, tooth- and bone-samples were investigated. Furthermore the mtDNA SNP analysis of a forensic case sample showing a mixed stain profile is presented. It was possible to discriminate two different haplogroups in this mixed-person stain. If compared to another mtDNA SNP profile that was found in a hair, the discriminating SNPs of the hair were as well found in the mixed-person stain.To disburden the SNP analysis in forensic casework, haplogroup assignment criteria and quality criteria for mtDNA SNaPshot analysis are announced.     

Genetic variation of 23 STR loci in a Northeast Colombian population (department of Santander)

The most efficient markers to solve filiation cases are the STRs, including complex cases that require the analysis of a greater number of markers. In this study samples from 123 unrelated individuals from the department of Santander (northeast Colombia) were typed for 23 autosomal STRs included in VeriFiler express kit (Thermo Fisher Scientific),and their allele frequencies and parameters of forensic relevance were determined. Results demonstrate independence within and between the loci analyzed, and the accumulate power of exclusion for the full set of markers was high (99.9996%), as well as the match probability, which was 1 in 8.77E + 29. Therefore, this northeast Colombian population database can be used in forensic to estimate the frequency of the genetic profile using of a multiple locus including in this DNA kit.     

4个STR基因座在广东汉族人群的单倍型分布

调查DYS19、DYS390和DYS389Ⅰ/Ⅱ基因座在广东汉族群体的单倍型分布。PCR扩增后,用不连续PAGE电泳系统分型。在130名无关男性个体中,共检出81种不同的单倍型,其中52种只出现过1次。4个基因座单倍型的遗传多样性、个体识别率和非父排除率分别为0.9989,0.9824,0.9824,结果表明,4个STR基因座的单倍型检测在法医学个体识别和亲权鉴定方面有较高的应用价值。     

Forensic and population genetic analysis of Serbian population using 21 STR loci of GlobalFiler™ PCR amplification kit

Autosomal short tandem repeats (STRs) have been widely used in forensic investigations. Prior to the application of any DNA based identification method, it is essential to estimate the allele frequencies and forensic statistical parameters of targeted STR loci in each population in order to provide a more precise reference database for forensic investigation. The GlobalFiler™ Kit is a multiplex assay that combines the 13 original CODIS loci with 7 non-overlapping loci from the expanded European Standard Set (ESS), as well as the highly discriminating SE33 locus, two Y-based loci and the sex determining maker, Amelogenin. The full complement of loci in the GlobalFiler™ Kit are: D13S317, D7S820, D5S818, CSF1PO, D1S1656, D12S391, D2S441, D10S1248, D18S51, FGA, D21S11, D8S1179, vWA, D16S539, TH01, D3S1358, AMEL, D2S1338, D19S433, DYS391, TPOX, D22S1045, SE33 and a Y-specific insertion/deletion locus (Yindel). The 6-dye GlobalFiler™ PCR Amplification kit (ThermoFisher Scientific) comprises 21 autosomal STRs have already been proven to be able to provide reliable DNA profiling results and enhance the power of discrimination between individuals. In this study, we are presenting an analysis of GlobalFiler STR loci on 209 unrelated individuals from Serbia.     

Canine Population Data Generated from a Multiplex STR Kit for Use in Forensic Casework*

Abstract: Canine biological specimens are often part of the physical evidence from crime scenes. Until now, there have been no validated canine‐specific forensic reagent kits available. A multiplex genotyping system, comprising 18 short tandem repeats (STRs) and a sex‐linked zinc finger locus for gender determination, was developed for generating population genetic data assessing the weight of canine forensic DNA profiles. Allele frequencies were estimated for 236 pedigreed and 431 mixed breed dogs residing in the U.S. Average random match probability is 1 in 2 × 10³³ using the regional database and 1 in 4 × 10³⁹ using the breed dataset. Each pedigreed population was genetically distinct and could be differentiated from the mixed breed dog population but genetic variation was not significantly correlated with geographic transition. Results herein support the use of the allele frequency data with the canine STR multiplex for conveying the significance of identity testing for forensic casework, parentage testing, and breed assignments.     

Chelex-100提取生物检材DNA实时PCR定量研究

目的研究Chelex-100法提取的生物检材DNA用量与复合STR分型成功率的关系。方法113份各种生物检材采用Chelex-100法提取DNA，应用Quantifiler人类DNA定量试剂盒在ABI 7500荧光定量PCR仪上进行实时PCR定量，同时用Identifiler复合扩增系统在ABI 3100遗传分析仪上对这些DNA样品进行STR分型。结果各种生物检材提取的DNA浓度分别为：37份滤纸、纱布血痕0．042～5．28ng／μl，16份口腔拭子1．15—4．21ng／μl，18份烟头0．016～1．46ng／μl，10份肋软骨0．531—14．40ng／μl，8份肌肉5．75—24．80ng／μl，7份指甲0．788—11．50ng／μl，17份精斑0．79～99．50ng／μl。在建立的8μl扩增体系中，根据上述结果，调整用于复合STR扩增的DNA模板量在0．5—3ng之间，大部分样品可获得完全的STR分型。结论Chelex-100法提取的检材DNA模板用量在0．5—3ng之间可得到有效STR扩增，浓度为0．5ng／μl以上的DNA样品，用小体积模板（1μl）比大体积（3μl）模板扩增效果好。     

Analysis of forensic samples in Banco Nacional de Datos Genéticos

Analysis of forensic samples to evaluate the rate of success for molecular markers: autosomal STRs, Y chromosome, and mitochondrial DNA. Since 2006 to date a total of 390 forensic samples were analyzed: bones, teeth, hairs, swabs, stains and paraffin embedded tissue. Bones and teeth, were pulverized in a Freezer Mill, extracted by chloroform/phenol/isoamyl alcohol method, and then purified with Centricon 100 columns. DNA from paraffin was extracted with QIAmp DNA Mini kit (QIAGEN). Mitochondrial DNA Control Region sequences were determined for regions HV1/HV2. Sequencing was performed using the BigDye^® Terminator v 1.1 Kit and analyzed in ABIPRISM^® 3100 Genetic Analyzer (AB). STRs were amplified using Amp FlSTR Identifiler^®, Minifiler^® and YFiler^® Kit (AB) and analyzed in ABI PRISM^® 3100 Genetic Analyzer and ABI PRISM^® 3130xl Genetic Analyzer (AB). Among forensic samples, bones and teeth analyzed for autosomal STRs, we obtained successful results in all of them. Incomplete typing are represented by loci of higher molecular weight, which demonstrates the poor quality of the sample due to its state of degradation and obtained better results using mini STRs. Successful results in sequencing for mitochondrial HV1 region for all samples analyzed, but in few hair samples we obtained mixed sequences and that represented important difficulties for the analysis. Age of samples and conservation are factors related which affect DNA viability. Autosomal STRs solved all the samples analyzed in our study, but Y chromosome analysis and mitochondrial DNA sequencing are also important and necessary markers in some forensic cases.     

Development of a Nomenclature System for a Canine STR Multiplex Reagent Kit*†

Abstract: Despite the popularity of dogs in US households, canine DNA evidence remains largely untapped in forensic investigations partially because of the absence of well‐defined forensic short tandem repeats (STRs), lack of standardized and validated PCR protocols, STR reagent kits, and poorly developed nomenclature. A nomenclature system was established based on internationally recognized recommendations for human forensic STRs for a recently developed canine STR reagent kit. Representative alleles were sequenced from each of the 18 STRs and the sex‐typing marker included in the kit. This study also reflects on the impact of point mutations, insertions, and deletions within and outside the STR core repeat structures. An understanding of the STRs’ sequence and repeat structures will enable development of a robust and reliable allele nomenclature and improve the accuracy and precision of allele fragment sizing in canine forensic profiling. The expected allele sizes have been calculated, and their repeat stuctures defined based on sequence information.     

Development of a real-time method to detect DNA degradation in forensic samples

Abstract: Knowledge of the degradation state of evidentiary DNA samples would allow selection of the appropriate analysis method (standard short tandem repeats [STRs] vs. mini STRs vs. mtDNA). This article describes the development of a Plexor® technology/real‐time PCR DNA degradation detection assay, which uses a common forward primer and two reverse primers (different fluorophores) to generate two Alu amplicons (63 and 246 bp). This very sensitive assay was optimized for reaction volume, cycle number, anneal/extend time, and temperature. Using DNA samples degraded with DNaseI, the ratio of the concentration of the short amplicon to the concentration of the long amplicon (degradation ratio) was increased versus time of degradation. Experiments were performed on a variety of environmentally degraded samples (age, sunlight, heat) and with seven commonly encountered forensic inhibitors. The degradation ratio was found to predict the observed loss of larger STR loci seen in the analysis of comprised samples.