Two examples of null alleles at the D19S433 locus due to the same 4 bp deletion in the presumptive primer binding site of the AmpFlSTR Identifiler kit

Two cases of allelic loss at the D19S433 locus after multiplex PCR with the AmpFlSTR Identifiler kit (Applied Biosystems) are described. In both cases the failure of PCR resulted in genetic inconsistencies due to opposite homozygosity. After singleplex PCR with published primers additional alleles were observed and Mendelian inheritance was restored. These PCR products were sequenced and in both cases the same 4 bp deletion near the 3′ end of the repeat region was detected in two alleles of different length. The frequency of these null alleles (two events in 1026 allelic transfers) amounts to 0.0019 (95% confidence limits: 0.0002-0.0070).     

Molecular characterization of a null allele at locus DXS8378

Typing of X-chromosomal short tandem repeat (STR) loci in a deficiency paternity case revealed a single Mendelian incompatibility between a female child and her putative grandmother, consisting of an opposite homozygosity at locus DXS8378. The presence of a null allele due to a primer binding site mutation on the child's paternally inherited X chromosome was confirmed by amplification with newly designed DXS8378 external primers. Sequencing analysis showed a point mutation (C > T transition at position 168, according to GenBank accession G08098) in the binding site of the original DXS8378 reverse primer.     

降解生物检材遗传标记检测的研究进展

降解生物检材遗传标记的检测一直是法医学实践和科研的难点。虽然PCR-STR分析技术能在一定程度上检测降解生物检材的遗传标记,但是对于某些高度降解的生物检材如骨骼、牙齿等的PCR-STR分型仍旧十分困难,存在不稳定性和分型异常现象。本文就近几年来PCR-STR分型技术在降解生物检材的法医实践中出现的问题和解决方法作一综述。     

Genetic polymorphism at 15 tetrameric short tandem repeat loci in four aboriginal tribal populations of Bengal

POPULATIONS: This study reports the genetic polymorphism observed at 15 short tandem repeat loci D3S1358, TH01, D21S11, D18S51, D5S818, D13S317, D7S820, D16S539, CSF1PO, vWA, D8S1179, TPOX, D2S1338, D19S433, and FGA in four aboriginal populations of Bengal. The analysis was performed to decipher the suitability of CODIS as well as six other highly polymorphic and unlinked markers in Forensic Testing. Studied populations include four tribes: Karmali, Kora, Maheli, and Lodha.     

A 21‐Locus Autosomal SNP Multiplex and its Application in Forensic Science

To develop a cost‐effective technique for single‐nucleotide polymorphism (SNP) genotyping and improve the efficiency to analyze degraded DNA, we have established a novel multiplex system including 21‐locus autosomal SNPs and amelogenin locus, which was based on allele‐specific amplification (ASA) and universal reporter primers (URP). The target amplicons for each of the 21 SNPs arranged from 63 base pair (bp) to 192 bp. The system was tested in 539 samples from three ethnic groups (Han, Mongolian, and Zhuang population) in China, and the total power of discrimination (TPD) and cumulative probability of exclusion (CPE) were more than 0.99999999 and 0.98, respectively. The system was further validated with forensic samples and full profiles could be achieved from degraded DNA and 63 case‐type samples. In summary, the multiplex system offers an effective technique for individual identification of forensic samples and is much more efficient in the analysis of degraded DNA compared with standard STR typing.     

Null allele can bring to interpretative problems in a deficitary paternity case

An STR null allele is an allele at a microsatellite locus that fails to amplify. A possible cause is poor primer annealing due to nucleotide sequence divergence in the flanking primers. In this study, a woman (ZAM) wanted to know whether a man (PGAF) was the father of her child (ZGC). During the court settlement, PGAF died. PGAF’s parents refused to undergo DNA investigation and denied the access to biological fragments from their dead son. Although, DNA specimens were obtained from buccal swabs of ZAM, ZGC and PGAF’s paternal sister (PTFS). Initially, only autosomal profiles were studied, and kinship assignment was inconclusive. Following our requests, PGAF’s parents (PRGF and LLGM) led us to obtain their DNA specimens. Only with the PTFS genetic profile, we were not able to demonstrate a kinship assignment. PTFS showed a homozygosis at D8S1179 locus. Then, merely comparing PTFS, LLGM and ZGC autosomal genetic profiles it was possible to underline that they were three different homozygous at D8S1179 locus. Hence, comparing the peak heights in different loci and according to literature, they had to carry a null allele at this locus. Parental studies were completed by Y haplotype analysis.     

Automated estimation of the number of contributors in autosomal STR profiles

Estimating the number of contributors to mixed STR profiles can be complex. This study describes the nC-tool to assist DNA expert in this process. The nC-tool is based on the total allele count for PowerPlex® Fusion 6C profiles and showed improved performance when compared to the maximum allele count approach.     

Inferring the Number of Contributors to Complex DNA Mixtures Using Three Methods: Exploring the Limits of Low‐Template DNA Interpretation

In forensic DNA casework, the interpretation of an evidentiary profile may be dependent upon the assumption on the number of individuals from whom the evidence arose. Three methods of inferring the number of contributors—NOCIt, maximum likelihood estimator, and maximum allele count, were evaluated using 100 test samples consisting of one to five contributors and 0.5–0.016 ng template DNA amplified with Identifiler^® Plus and PowerPlex^® 16 HS. Results indicate that NOCIt was the most accurate method of the three, requiring 0.07 ng template DNA from any one contributor to consistently estimate the true number of contributors. Additionally, NOCIt returned repeatable results for 91% of samples analyzed in quintuplicate, while 50 single‐source standards proved sufficient to calibrate the software. The data indicate that computational methods that employ a quantitative, probabilistic approach provide improved accuracy and additional pertinent information such as the uncertainty associated with the inferred number of contributors.     

The null hypothesis is not called that for nothing: statistical tests in randomized trials

This article aims to update readers on different ways to arrange one’s thinking about conventional null hypotheses in randomized trials. It covers basic criticism of conventional hypotheses and, beyond this, covers relevant developments in methodological, organizational, and science policy arenas. This article includes coverage of new ways to frame null hypotheses, new technical resources, standards for registering trials and reporting on them, cumulating results, common mistakes, and post-trial analysis of null results. The paper includes ideas for research and development on each topic.

Powerplex® Y 23 System: Molecular characterization of a null allele at locus DYS549

We observed a null allele pattern at locus DYS549 in a male subject from North-East Italy typed with the PowerPlex^® Y 23 System (Promega). To investigate whether this pattern was due to the presence of a microdeletion/mutation in primer binding sites or in the locus target region, the sample was amplified with our designed DYS549 primers obtained from GenBank sequence (GDB: 515022). After amplification, a normal hemizygous genotype at this locus was generated, thus indicating the presence of a point mutation in the binding site of the original primer set of PowerPlex^® Y 23 System (Promega). This was further confirmed by sequence analysis, carried out with the Big Dye Terminator v3.1 Cycle Sequencing kit (Applied Biosystems), according to the manufacturer's instructions. Sequences were run on the ABI Prism 3130 Genetic Analyzer (Applied Biosystems) and analyzed using the Sequencing Analysis v.5.3.1 and the SeqScape v2.6 softwares (Applied Biosystems). Ascertainment of the frequency of null alleles generated from variations at primer binding sites of short tandem repeats loci is of great importance in forensic genetics.