Methods for ensuring the highest DNA concentration and yield in future and retrospective trace DNA extracts

In forensic laboratories, increased extraction efficiency of trace evidence is paramount because analytical success is intrinsically dependent on the quantity of DNA recovered. Moreover, highly concentrated nucleic acids are vital for effective downstream analysis and high quality results. This study investigated the efficiency of extraction with the Qiagen® QIAamp® DNA Investigator kit, and explored improvements to the methodology that would maximise the recovery of low concentration forensic samples. Controlled amounts of starting cellular material were used to mimic trace (or low level) DNA deposits prior to DNA extraction with the Investigator kit. Addition of the provided carrier RNA along with conducting two successive elutions of 50 µL improved the net recovery of DNA to 95%. Concentration with centrifugal filters post-extraction were able to concentrate DNA but a large net loss was observed. For the concentration of historic, retrospectively extracted DNA, centrifugal methods are able to concentrate DNA extracts previously too dilute for analysis. These concentrated volumes, however are small, allowing for minimal downstream analysis attempts before the sample is exhausted.     

Blood and sperm traces on human hair. A study on preservation and detection after 3-month outdoor exposure

Hair can retain important biological traces for forensic investigations. Forensic scientists are used to looking for such traces on clothing and skin of victims, however, when decomposition kicks in and all that remains of the victims is the skeleton, hair may be the only tissue representing the surface of the body at the time of a crime on which biological traces of an aggressor may have been left and still be detectable. Given the lack of research on this topic, this pilot study aims to assess the capacity of hair to retain semen and blood in hair, and the possibility to detect these fluids with well-known techniques and to obtain a useful genetic profile even when exposed to environmental conditions (Open Natural Environment (woods), Open Man Made Environment (urban)) for three months.Results showed that both traces were always visible and detectable with almost all techniques in the Control Environment, while in the two open environments some difficulties arose. However, biomolecular analysis was effective up to three months on both fluids in the Natural Environment and up to two months and one week respectively on blood and semen in the Man Made Environment. The Combur Test, OBTI, and Luminol were effective on blood up to three months in both environments while Sperm-HY-Liter and observation of cellular components were effective on semen up to at least 1 month and PSA testing was positive up to 1 week in both environments.The present work can be considered an encouraging starting point for the analysis of biological traces on hair in forensic contexts, regardless of the PMI, since blood and semen related to a crime may survive.     

基于年龄相关DNA甲基化位点推断河南汉族个体年龄

目的探查中国河南汉族个体与年龄相关的DNA甲基化位点,构建年龄推断模型,进行甲基化和年龄相关性分析。方法采用焦磷酸测序法对ELOVL2、ClOrf132、KLF14、TRIM59和FHL2基因的34个CG位点进行甲基化分析,利用SPSS 23软件的多元回归方法建立模型,对甲基化和年龄相关性做分析。结果除ClOrf132基因3个CG位点的甲基化水平与年龄呈负相关外,其余4个基因的31个CG位点甲基化水平均与年龄呈正相关。多元回归分析表明,年龄与CG位点的甲基化水平存在明显的线性关系,实际年龄与推断年龄偏差在5岁以内的准确度达80%以上。结论本研究构建的河南汉族个体年龄推断模型,有助于通过检测血液等组织的DNA甲基化水平推断个体的年龄范围,具有法医学应用前景。     

State as resource,mediator and performer: understanding the local and global politics of gold mining in Kyrgyzstan

The inability of the state to maintain security and the rule of law for the purposes of foreign direct investment and industrial production is often taken as a sign of its weakness. However, such judgments say little about the actual functions of the state for global extraction industries and local political forces which demand their share of the pie. Whilst coercive state power may have decreased since Kyrgyzstan became independent, more important is the fact that the state itself has been transformed under the ruptures of, on the one hand, economic and political liberalization and, on the other, the effects of so-called ‘revolutions' of 2005 and 2010 which led to the wholesale restructuring of national structures of clientelism. Based on ethnographic research conducted in Talas province, documentary sources and interviews with gold mining companies and state officials, the paper investigates the state's shifting roles with respect to Kyrgyzstan's gold mining sector. Firstly, it explores the state as a source of rents for officials who grant and rescind licences in exchange for formal and informal payments from foreign investors, often via offshore vehicles. Secondly, it considers the role of the state as mediator between foreign investors and their access to sites. Finally, it identifies the state as performer of its status as sovereign power despite its inability to prevent uprisings and actually guarantee the promised access to its territory.     

Nondestructive Methods for Recovery of Biological Material from Human Teeth for DNA Extraction

The extraction of DNA from human skeletal remains applied to forensic, and evolutionary studies do not exclude risks, which are to be evaluated when working with unique specimens that could be damaged or even destroyed. In the present study were evaluated several nondestructive methods for recovering DNA instead of the most currently used pulverization method. Three different procedures to access inside the dental pieces (occlusal perforation, cervical perforation, and cervical cut) have been compared with the aim of recovering as many cell remains as possible to carry out a DNA extraction. Given the DNA quantitation results, a method was proposed that consists of a cervical cut to facilitate the access to the pulp cavity and a subsequent filing of the root canals down to the apex of the dental root. This methodology allows the recovery of both mitochondrial and nuclear DNA, with the minimum deterioration for the dental pieces.     

Evaluation of a Visualization Assay for Blood on Forensic Evidence

In forensics, bloodstains on dark fabrics might be invisible for the naked eye. Although several visualization, presumptive, and confirmatory blood tests have been developed, all have one or more disadvantages, especially on DNA analysis. We report here the use of a visualization assay that can visually detect blood drops up to 1/20 dilution. In this assay, the fabric is placed between two wet filter papers and covered by glass surfaces on both sides. Pressure is applied on the glass surfaces in which bloodstains transfer onto the filter papers through capillary forces. Detected stains can be tested with other more sensitive presumptive blood tests performed on the filter paper. Even more, DNA analysis can be performed on the transferred bloodstains. The presented visualization assay is easy to perform, extremely cheap, requires little hands on time, and does not affect bloodstain pattern analysis.     

A Simple Digestion Method with a Lefort Aqua Regia Solution for Diatom Extraction

Presence of diatoms in tissues has been considered as a significant sign of drowning. However, there are limitations in the present extraction methods. We developed a new digestion method using the Lefort aqua regia solution (3:1 nitric acid to hydrochloric acid) for diatom extraction and evaluated the digestive capability, diatom destruction, and diatoms' recovery of this new method. The kidney tissues from rabbit mixed with water rich in diatoms were treated by the Lefort aqua regia digestion method (n = 10) and the conventional acid digestion method (n = 10). The results showed that the digestive capability of Lefort aqua regia digestion method was superior to conventional acid digestion method (p < 0.01); the structure of diatom remained almost intact; and the recovery of diatom was comparable to the conventional acid digestion method (p > 0.05). The Lefort aqua regia reagent is an improvement over the conventional acid digestion for recovery of diatoms from tissue samples.     

TrueAllele® Genotype Identification on DNA Mixtures Containing up to Five Unknown Contributors

Computer methods have been developed for mathematically interpreting mixed and low‐template DNA. The genotype modeling approach computationally separates out the contributors to a mixture, with uncertainty represented through probability. Comparison of inferred genotypes calculates a likelihood ratio (LR), which measures identification information. This study statistically examined the genotype modeling performance of Cybergenetics TrueAllele^® computer system. High‐ and low‐template DNA mixtures of known randomized composition containing 2, 3, 4, and 5 contributors were tested. Sensitivity, specificity, and reproducibility were established through LR quantification in each of these eight groups. Covariance analysis found LR behavior to be relatively invariant to DNA amount or contributor number. Analysis of variance found that consistent solutions were produced, once a sufficient number of contributors were considered. This study demonstrates the reliability of TrueAllele interpretation on complex DNA mixtures of representative casework composition. The results can help predict an information outcome for a DNA mixture analysis.     

An Accelerated Analytical Process for the Development of STR Profiles for Casework Samples

Significant efforts are being devoted to the development of methods enabling rapid generation of short tandem repeat (STR) profiles in order to reduce turnaround times for the delivery of human identification results from biological evidence. Some of the proposed solutions are still costly and low throughput. This study describes the optimization of an analytical process enabling the generation of complete STR profiles (single‐source or mixed profiles) for human identification in approximately 5 h. This accelerated process uses currently available reagents and standard laboratory equipment. It includes a 30‐min lysis step, a 27‐min DNA extraction using the Promega Maxwell^®16 System, DNA quantification in <1 h using the Qiagen Investigator^® Quantiplex HYres kit, fast amplification (<26 min) of the loci included in AmpF?STR^® Identifiler^®, and analysis of the profiles on the 3500‐series Genetic Analyzer. This combination of fast individual steps produces high‐quality profiling results and offers a cost‐effective alternative approach to rapid DNA analysis.     

温肾养血方对无排卵模型大鼠卵巢质量和形态的影响

目的 探讨温肾养血方对无排卵大鼠子宫指数、卵巢指数及卵巢组织形态的影响。方法 采用丙酸睾酮皮下注射法复制无排卵大鼠模型。将模型大鼠分为模型组,西药组,温肾养血方大、中、小剂量组,分别给予生理盐水,枸橼酸氯米芬,温肾养血方大剂量（52 mg/kg）、中剂量（26 mg/kg）、小剂量（13 mg/kg）灌胃。另选22日龄大鼠作为正常组。连续灌胃21 d后,测定各组大鼠体质量、子宫指数和卵巢指数,光镜下观察卵巢组织形态。结果 与正常组比较,模型组大鼠体质量无明显变化（P>0.05）,子宫指数和卵巢指数显著降低（P<0.05）。与模型组比较,各给药组大鼠体质量和子宫指数、卵巢指数无明显变化（P>0.05）。温肾养血方大、中剂量组卵巢形态近似正常组,结构明显改善,多数卵巢可见发育期的各级卵泡,可见卵丘、卵细胞及典型黄体形成。西药组和温肾养血方小剂量组卵巢形态类似模型组,卵巢体积较小,卵巢表面苍白,无斑点,色泽苍白,卵泡数目少,卵泡较小,且形状不规则,多呈囊性扩张,无黄体。结论 温肾养血方能改善卵巢结构和促进卵泡发育。