STR基因座荧光标记复合扩增检测及其法医学应用

目的建立荧光标记复合扩增检测4个STR基因座的方法,应用于法医学亲权鉴定与个人识别。方法用6-FAM标记D3S1754引物,HEX标记D4S2366和D12S375引物,TMR标记D1S549引物,PCR复合扩增,310基因分析仪电泳自动收集电泳结果数据,GeneScanAnalysisSoftware3.7NT计算扩增产物片段相对大小,Genotyper誖3.7NT软件进行样本基因型分型。将该方法应用于法医学亲权鉴定与个人识别。结果建立了荧光标记复合扩增检测4个STR基因座基因型的方法,具有良好的稳定性和较好的灵敏度,分型结果准确。结论研究建立的方法操作简便,分型结果稳定可靠,可用于遗传多态性研究和法医学亲权鉴定与个人识别。     

Population data for four population groups from the United States for the eleven Y-chromosome STR loci recommended by SWGDAM

POPULATIONS: Caucasian ( n =1243), African American ( n =1605), Hispanic ( n =454), and Native American ( n =104).     

Simultaneous determination of total human and male DNA using a duplex real-time PCR assay

A single duplex assay to determine both the amount of total human DNA and the amount of male DNA in a forensic sample has been developed. This assay is based on TaqMan technology and uses the multicopy Alu sequence to quantitate total human DNA and the multicopy DYZ5 sequence to quantitate Y chromosomal (male) DNA. The assay accepts a wide concentration range of input DNA (2 muL of 64 ng/microL to 0.5 pg/microL), and also allows detection of PCR failure. The PCR product sizes Alu (127 bp) and DYZ5 (137bp) approximate that of the smaller short tandem repeats (STRs) which should make the assay predictive of STR success with degraded DNA. The assay was optimized for probe/primer concentrations and BSA addition and validated on its reproducibility, on its human specificity, on its nonethnic variability, for artificial mixtures and adjudicated casework, for the effect of inhibitors and for state of DNA degradation. This assay should prove very usual in forensic analyses because knowing the relative amounts of male versus female DNA can allow the examiner to decide which samples may yield the most probative value in a case or direct the samples to methods that would yield the greatest information.     

Age Estimation in Living Egyptians Using Signal Joint T‐cell Receptor Excision Circle Rearrangement

Age estimation is one of the challenges in forensic sciences. There are many techniques to estimate the age. Molecular biology approach is one of these techniques. Signal joint T‐cell receptor excision circles gene (sjTRECs), is one of this approach. We aimed to use sjTRECs as a suitable marker for age estimation among Egyptian population. TaqMan qPCR approach was used to quantify sjTREC levels in blood samples obtained from 153 healthy Egyptian individuals ranging from a few weeks to 70 years. Our results showed a significant negative correlation between sjTREC levels and age with p ≤ 0.05. Moreover, the individual's age can be determined through this formula Age = ?30.671+ (?5.998Y) (Y is dCtTBP ? sjTREC) with standard error ±7.35 years. Within the forensic context, sjTREC' levels can be used to estimate the Egyptian individual's age accurately.     

Quantifiler observations of relevance to forensic casework

The Quantifiler (QF) kit is regularly used by forensic scientists for DNA quantitation. We performed in-house validation studies which revealed some interesting observations. The QF standard displayed a two-fold difference between two different lot numbers which suggests that every standard should be tested prior to use. The Promega K562 DNA standard works well with the QF kit. c. 41% of samples that inhibited the internal PCR control (IPC) system within the QF kit still produced good Profiler Plus reactions. QIAquick was effective at removing inhibitors. The presence of dyes within casework samples were observed not to inhibit QF amplifications. Template DNA greater than 100 ng/muL appeared to inhibit the IPC. Close to identical concentration results were obtained when alternative analysis settings were used. These validation findings will assist DNA processes involved in forensic casework.     

Direct STR amplification from whole blood and blood- or saliva-spotted FTA without DNA purification

The DNA purification step has been thought to be essential for typing of STR DNA. However, this process is time-consuming, and there is a risk of unexpected cross-contamination during purification. We report a new method for direct short tandem repeat (STR) amplification using a newly developed direct PCR buffer, AnyDirect, which can amplify STR loci from whole blood and blood- or saliva-spotted FTA cards without DNA purification. The autosomal and Y chromosomal STR loci were analyzed for whole blood and blood or saliva spots of random individuals, followed by comparison of the results with those of corresponding purified DNA. The results from whole blood and blood spots showed perfect concordance with those from purified DNA without allele or locus drop-out. However, in the case of saliva spots, no amplification or locus drop-out was observed in some of the samples, which offers a topic for further study. Additionally, some commercial hot-start DNA polymerases other than AmpliTaq Gold DNA polymerase were also found to be compatible with this buffer system. Therefore, this direct PCR buffer was demonstrated to be useful for fast forensic DNA analysis or criminal DNA databases for which there is no need to store DNA samples.