Interrogation of short tandem repeats using fluorescent probes and melting curve analysis: A step towards rapid DNA identity screening

Current forensic DNA profiling methods rely on the analysis of samples at specialised laboratories with an average turnaround time of several days. The ability to rapidly determine a partial profile of short tandem repeats at the point-of-arrest would be of great benefit to police forces around the world, for example enabling a suspect to be rapidly included or excluded from an investigation. We have developed a homogeneous PCR method for the interrogation of STR loci utilising fluorescent oligonucleotide probes and melting curve analysis. Alleles of the D18S51, TH01 and D8S1179 loci were differentiated and identified on the basis of target length and probe melting temperature. Assay performance was evaluated by comparing melting peak data with the AmpFlSTR^® SGM Plus^® system. The method is compatible with direct analysis of unpurified buccal swab samples, enabling a partial STR profile to be generated within 1 h.     

Polymorphism of eight X-chromosomal STRs in a Japanese population

Eight X-chromosomal short tandem repeat (X-STR) markers were analyzed in 258 unrelated Japanese (144 males and 114 females) using Mentype^® Argus X-8 PCR Amplification Kit (Biotype AG) which contains DXS7132, DXS7423, DXS8378, DXS10074, DXS10101, DXS10134, DXS10135 and HPRTB. The DXS10135 locus proved to be highly polymorphic marker (PIC: 0.945) and the DXS7423 showed the lowest value (PIC: 0.453). The exact test for genotype distribution showed no significant deviation from the Hardy-Weinberg equilibrium.     

Population data of eight X-chromosomal STR markers in Ewe individuals from Ghana

The investigation of the X-linked DNA markers are well established in the forensic routine case work. We studied an Ewe population sample from Ghana. The eight X-chromosomal STRs DXS10135, DXS8378, DXS7132, DXS10074, HPRTB, DXS10101, DXS10134 and DXS7423 were analyzed in 182 Ewe individuals (108 females and 74 males) from the region of Sogakofe (Ghana). Allele frequencies and statistical parameter as well as comparison with known data from Germans and with data from an Amharic population (Ethiopia) are presented.     

Post-injection hybridization of complementary DNA strands on capillary electrophoresis platforms: A novel solution for dsDNA artifacts

Several laboratories have reported the occurrence of a split or n − 1 peak at the vWA locus in PowerPlex^® 16 and PowerPlex^® ES amplification products separated on 4- and 16-capillary electrophoresis instruments. The root cause of this artifact is post-PCR reannealing of the unlabeled, unincorporated vWA primer to the 3′-end of the tetramethylrhodamine (TMR)-labeled strand of the vWA amplicon. This reannealing occurs in the capillary post-electrokinetic injection. The split peak is eliminated by incorporation into the loading cocktail of a sacrificial hybridization sequence (SHS) oligonucleotide that is complementary to the vWA primer. The SHS preferentially anneals to the primer instead of the TMR-labeled strand of the vWA amplicon. In addition, the n − 10/n − 18 artifact that may be seen at the vWA locus was determined to be due to double-stranded amplicon formed post-electrokinetic injection into the capillary. This was also eliminated by adding in two Complementary Oligo Targets (COT1 and COT2) in addition to the SHS oligonucleotide into the loading cocktail. These three oligonucleotides are complementary to the 33 bases at the 5′-end of the unlabeled vWA amplicon strand and the 60 bases at its 3′-end and therefore compete for hybridization to the TMR-labeled amplicon strand. Incorporation of these three oligonucleotides in the Internal Lane Standard 600 (ILS600) eliminate both the split peak and n − 10/n − 18 artifact in PowerPlex^® 16 and PowerPlex^® ES amplification products without affecting sizing of alleles at the vWA locus or any locus in the PowerPlex^® 16, PowerPlex^® Y, PowerPlex^® ES, AmpFlSTR^® Profiler Plus^® ID, AmpFlSTR^® Cofiler^®, and AmpFlSTR^® SGM Plus^® kits.     

An ambiguous sequence-based allele of SE33

SE33 was a well-known autosomal short tandem repeat (STR) marker that was high polymorphic and therefore was high discrimination power. The sequence structure of STR markers has been increasingly explored with next-generation sequencing (NGS) technology. The sequencing resulted in the development of a new locus designation and allele nomenclature that was also backward compatible with the conventional capillary electrophoresis. SE33 was one of the STR markers that had been coamplified by Forenseq™ Signature Prep Kit (Verogen) but were not analyzed and illustrated in the Universal Analysis Software (UAS) (Verogen). This study reported an ambiguous sequence-based allele 16.3 of the SE33 locus. This allele was observed while analyzed by STRait Razor 3.0. The configuration file was modified from the previous studies to include 15 bp of 5′ flanking region and 24 bp of 3′ flanking region. The ambiguous allele was called 16.3 (106 bp) with a read count of 2070. However, the sequence of the repeat region cannot be designated as allele 16.3. Several possible scenarios for allele designation were presented and discussed.     

用串联质谱法提高毒鼠强的检测灵敏度

目的建立一种能避免生物样品中杂质的干扰,提高毒鼠强检测灵敏度的方法。方法应用串联质谱法,对生物样品中的毒鼠强进行检验研究。结果在样品无需净化的情况下,以m/z240为母离子,当激活电压在0.8V左右时,毒鼠强二次电离的离子碎片适中,质谱谱图清晰;此时进样量为1.0ng的毒鼠强,其信噪比(s/n)为87,经与全扫描质谱法进行比较灵敏度大为提高。结论该方法能有效提高毒鼠强检测灵敏度,适用于同类案件检验。     

D18S872基因座在汉、维、蒙古、回族群体中的遗传多态性研究及其应用

目的　调查D18S872基因座在成都汉族 ,新疆维族和蒙古族 ,甘肃回族 4个民族中的遗传多态性 ,获得群体遗传学基本数据。　方法　等位基因分型标准物制备采用分子克隆技术 ,样本基因分型采用PCR和PAG垂直电泳技术、银染显色方法。　结果　获得D18S872基因座等位基因分型标准物及该基因座在 4个群体中的遗传学数据。　结论　结果表明D18S872基因座在法医学个人识别和亲子鉴定中有一定的应用价值。     

Allele frequencies of nine STR loci—D16S539, D7S820, D13S317, CSF1PO, TPOX, TH01, F13A01, FESFPS and vWA—in the population from Alagoas, northeastern Brazil

The polymorphism of nine STR loci has been studied in a sample of 598 individuals from the population of Alagoas, Northeastern Brazil. Determination of the allele frequencies as well as of several commonly used statistics in forensic and paternity testing were defined. The most polymorphic loci were TH01 and D7S317. The exact test demonstrated that the nine loci analyzed in the population have no deviation from Hardy–Weinberg equilibrium (P>0.05).     

生物组织中甲胺磷及其异构体的串联质谱法分析

目的建立可用于生物体内甲胺磷及其异构体O,O二甲基氨基硫代磷酸酯检验的高灵敏度方法。方法应用串联质谱法(GCMS/MS),对生物样品中的甲胺磷及其异构体进行检验研究。结果选择质荷比为141的离子为母离子,击发电压为0.8V时,甲胺磷及其异构体二次电离碎片适中,质谱谱图清晰。50%甲胺磷乳油进样量为0.1ng时,甲胺磷及O,O二甲基氨基硫代磷酸酯峰的信噪比分别为436、773。经实验,甲胺磷检测限为20pg。结论所拟方法能有效提高甲胺磷的检测灵敏度。     

15个基因座复合扩增体系的建立及新疆维吾尔族遗传多态性

目的建立15个基因座五色荧光复合扩增体系,并调查新疆维吾尔族的遗传多态性。方法筛选STR基因座,等位基因测序后按照重复序列重复次数命名,对建立的15个基因座五色荧光复合扩增体系进行灵敏度、种属特异性、同一性及稳定性检测,对新疆维吾尔族群体进行遗传多态性分析,与西藏藏族、岫岩满族、广州汉族进行群体间比较。结果建立了15个基因座复合扩增体系,检测灵敏度为0.3 ng,具有良好的种属特异性、同一性和稳定性。新疆维吾尔族13个常染色体STR基因座的基因频率分布符合Hardy-Weinberg平衡,无连锁不平衡现象,大部分基因座在群体间差异具有统计学意义。结论建立的体系具有法医物证学应用价值,13个常染色体STR基因座在新疆维吾尔族群体多态性高,适合于亲子鉴定及个人识别,可作为现有基因座的补充。