Investigations into the hypothesis of transgenic cannabis

The unusual concentration of cannabinoids recently found in marijuana samples submitted to the forensic laboratory for chemical analysis prompted an investigation into whether genetic modifications have been made to the DNA of Cannabis sativa L. to increase its potency. Traditional methods for the detection of genetically modified organisms (GMO) were used to analyze herbal cannabis preparations. Our analyses support the hypothesis that marijuana samples submitted to forensic laboratories and characterized by an abnormal level of Δ(9)-THC are the product of breeding selection rather than of transgenic modifications. Further, this research has shown a risk of false positive results associated with the poor quality of the seized samples and probably due to the contamination by other transgenic vegetable products. On the other hand, based on these data, a conclusive distinction between the hypothesis of GMO plant contamination and the other of genetic modification of cannabis cannot be made requiring further studies on comparative chemical and genetic analyses to find out an explanation for the recently detected increased potency of cannabis.     

A new pentaplex system to study short tandem repeat markers of forensic interest on X chromosome

A new method has been optimised to amplify five X chromosome short tandem repeat (STR) markers of interest in forensic medicine: human phosphoribosyl transferase (HPRTB), DXS101, androgen receptor (ARA), DXS7423 and DXS8377. Markers were conveniently amplified in a single PCR reaction with fluorochrome-labelled primers, which allowed the analysis of fragment sizes after injection into a capillary electrophoresis system. The most common alleles of each locus were sequenced and used in a control ladder to type unknown samples.     

Increasing Allele Detection by Altering the Quantity of Internal Lane Standard

Electrokinetic injection (EI) is the primary method used in forensic laboratories to load amplified PCR product in capillary electrophoresis for short tandem repeat (STR) fragment separation. Because all samples subjected to capillary electrophoresis use internal lane standard (ILS), this study investigated the consequence of varying the volume of ILS and its effects on allele peak heights and number of alleles detected. Results demonstrated that when the volume of ILS is reduced, the average peak height and number of alleles increased, thereby increasing the sensitivity of the detection method. Sizing anomalies were observed; however, they did not adversely affect accuracy and precision. The method developed in this study offers a simple and universal procedure to increase the alleles detected in forensic STR analysis. Reducing the volume of ILS to achieve greater sensitivity is applicable to all STR amplification kits and capillary electrophoresis instruments currently used in forensic DNA analysis.     

DNA Purification Cell Lysis and Wash Step Modifications for Low-Template DNA Sample Processing,

As DNA technology becomes increasingly sensitive, forensic laboratories are receiving more low-template DNA samples. These samples, already low in DNA content, become even more challenging to process as the available DNA becomes further reduced during the extraction step. In this study, two extraction modifications were tested to determine if the cause of DNA loss could be identified and mitigated. A double lysis technique was used to test for DNA loss in the sample collection substrate, and lysate eluates were re-extracted to determine DNA loss from inefficient binding to the silica column. Both modifications showed DNA was lost at these steps. However, resulting STR profiles from these samples had fewer peaks and lower peak heights when compared to samples processed with no extraction modifications. Overall, the potential benefits of adding these extraction modifications for low-template DNA sample processing are not enough to justify the risk associated with additional manipulation.     

焦磷酸测序技术分析单核苷酸多态性在法医学中的应用

单核苷酸多态性（SNP）是新一代的法医学遗传标记,有望成为法医实践中解决高度降解检材及特殊案件DNA鉴定的重要工具。近年来涌现出许多高通量的SNP分析方法,如引物延伸结合时间飞行质谱分析法、微测序法、SNP lex以及焦磷酸测序法等。本文重点对焦磷酸测序技术的原理、步骤及其在法医学中的应用进展进行简要的综述。     

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.     

Isoelectric focusing of human hair keratins: correlation with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and effect of cosmetic treatments.

A new isoelectric focusing (IEF) technique in polyacrylamide gels with 6M urea and 1.5% Nonidet P40 has been developed to characterize human hair samples. The phenotypes demonstrated with this procedure has been correlated with the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns described by other authors. The method described can be applied in the forensic science analysis of a single human hair. Using the same IEF technique we have studied the changes in electrophoretic patterns of cosmetically treated hair. The characteristics of the modifications observed and its utility in forensic science work are also discussed in this paper.     

英国法庭科学服务部的市场化变迁及其启示

英国法庭科学服务部市场化发展历史体现出竞争性市场有利于提升效率和为买方节约成本，这说明纯公益性刑事司法服务的某些法庭科学服务项目是可以市场化的；但其最终因超高的营运成本被迫关闭也说明商品化的服务市场承载不了法庭科学全面发展所需要的科学性要求。英国采用价格竞争的方式将法庭科学服务市场化，相比之下，真正的竞争性在中国当前的司法鉴定制度下却不存在，对司法鉴定制度进行改革已经刻不容缓。     

Application of the BioMek 2000 Laboratory Automation Workstation and the DNA IQ System to the extraction of forensic casework samples

Robotic systems are commonly utilized for the extraction of database samples. However, the application of robotic extraction to forensic casework samples is a more daunting task. Such a system must be versatile enough to accommodate a wide range of samples that may contain greatly varying amounts of DNA, but it must also pose no more risk of contamination than the manual DNA extraction methods. This study demonstrates that the BioMek 2000 Laboratory Automation Workstation, used in combination with the DNA IQ System, is versatile enough to accommodate the wide range of samples typically encountered by a crime laboratory. The use of a silica coated paramagnetic resin, as with the DNA IQ System, facilitates the adaptation of an open well, hands off, robotic system to the extraction of casework samples since no filtration or centrifugation steps are needed. Moreover, the DNA remains tightly coupled to the silica coated paramagnetic resin for the entire process until the elution step. A short pre-extraction incubation step is necessary prior to loading samples onto the robot and it is at this step that most modifications are made to accommodate the different sample types and substrates commonly encountered with forensic evidentiary samples. Sexual assault (mixed stain) samples, cigarette butts, blood stains, buccal swabs, and various tissue samples were successfully extracted with the BioMek 2000 Laboratory Automation Workstation and the DNA IQ System, with no evidence of contamination throughout the extensive validation studies reported here.     

Applicability of DNA analysis on adhesive tape in forensic casework

Adhesive tape is commonly used in crimes and is often the subject of forensic evaluation. DNA analysis of adhesive tape can provide DNA profiles of suspects. The object of this study was to evaluate the applicability of DNA analysis on adhesive tape samples in forensic casework. We retrospectively reviewed all cases involving adhesive tape or similar items received by our institute for DNA analysis during the past 11 years. From 100 forensic cases reviewed, 150 adhesive tape samples were examined. A total of 98 DNA profiles were obtained from these samples. Sixty-two of the profiles provided feasible case-relevant information. In conclusion, DNA profiling of adhesive tape samples can be useful in a variety of forensic cases.     

文件鉴定标准体系的研究

本文介绍了我国文件鉴定标准化建设的现状及其意义,并根据文件检验学学科和鉴定质量活动的特点,结合我们在文件鉴定实验室标准化建设实践中的经验,详细阐述了文件鉴定标准体系的结构,及文件鉴定系列标准(及部分)应包含的主要内容,希望能对我国文件鉴定标准化建设起到积极的推动作用。     

Considerations on the ASTM Standards 1789‐04 and 1422‐05 on the Forensic Examination of Ink

Abstract: The ASTM standards on Writing Ink Identification (ASTM 1789‐04) and on Writing Ink Comparison (ASTM 1422‐05) are the most up‐to‐date guidelines that have been published on the forensic analysis of ink. The aim of these documents is to cover most aspects of the forensic analysis of ink evidence, from the analysis of ink samples, the comparison of the analytical profile of these samples (with the aim to differentiate them or not), through to the interpretation of the result of the examination of these samples in a forensic context. Significant evolutions in the technology available to forensic scientists, in the quality assurance requirements brought onto them, and in the understanding of frameworks to interpret forensic evidence have been made in recent years. This article reviews the two standards in the light of these evolutions and proposes some practical improvements in terms of the standardization of the analyses, the comparison of ink samples, and the interpretation of ink examination. Some of these suggestions have already been included in a DHS funded project aimed at creating a digital ink library for the United States Secret Service.     

A DNA microarray system for forensic SNP analysis

Forensic DNA analysis is routinely performed using polymorphic short tandem repeat (STR) markers. However, for degraded or minute DNA samples, analysis of autosomal single nucleotide polymorphisms (SNPs) in short fragments might be more successful. Furthermore, sequencing of mitochondrial DNA (mtDNA) is often performed on highly degraded or scarce samples due to the high copy number of mtDNA in each cell. Due to the increasing number of complete mtDNA genome sequences available, the limited discrimination power of an mtDNA analysis, may be increased by analysis of coding region polymorphisms in addition to the non-coding variation. Since sequence analysis of the coding region would require more material than generally present in forensic samples, an alternative SNP analysis approach is required. We have developed a one-colour microarray-based SNP detection system for limited forensic materials. The method is based on minisequencing in solution prior to hybridisation to universal tag-arrays. In a first outline of a forensic chip, a combination of 12 nuclear and 21 mitochondrial SNP markers are analysed simultaneously. The mitochondrial markers on the chip are polymorphisms within the hypervariable region as well as in the coding region. Even though the number of markers in the current system is limited, it can easily be extended to yield a greater power of discrimination. When fully developed, microarray analysis provides a promising system for efficient sensitive SNP analysis of forensic samples in the future.     

Y-chromosome STR system, Y-PLEX 12, for forensic casework: development and validation

The Y-PLEX 12 system, developed for use in human identification, enables simultaneous amplification of eleven polymorphic short tandem repeat (STR) loci, namely DYS392, DYS390, DYS385 a/b, DYS393, DYS389I, DYS391, DYS389II, DYS 19, DYS439 and DYS438, residing on the Y chromosome and Amelogenin. Amelogenin provides results for gender identification and serves as internal control for PCR. The validation studies were performed according to the DNA Advisory Board's (DAB) Quality Assurance Standards. The minimal sensitivity of the Y-PLEX 12 system was 0.1 ng of male DNA. The mean stutter values ranged between 3.76-15.72%. A full male profile was observed in mixture samples containing 0.5 ng of male DNA and up to 400 ng of female DNA. Amelogenin did not adversely affect the amplification of Y-STRs in mixture samples containing male and female DNA. The primers for the Y-STR loci present in Y-PLEX 12 are specific for human DNA and some higher primates. None of the primate samples tested provided a complete profile at all 11 Y-STR loci amplified with the Y-PLEX 12 system. Y-PLEX 12 is a sensitive, valid, reliable, and robust multiplex system for forensic analysis, and it can be used in human forensic and male lineage identification cases.     

Developmental validation of a multiplex qPCR assay for assessing the quantity and quality of nuclear DNA in forensic samples

Forensic scientists are constantly searching for better, faster, and less expensive ways to increase the first-pass success rate of forensic sample analysis. Technological advances continue to increase the sensitivity of analysis methods to enable genotyping of samples containing minimal amounts of DNA, yet few tools are available that can simultaneously alert the analyst to both the presence of inhibition and level of degradation in samples prior to genotyping to allow analysts the opportunity to make appropriate modifications to their protocols and, consequently, to use less sample. Our laboratory developed a multiplex quantitative PCR assay that amplifies two human nuclear DNA target sequences of different length to assess DNA degradation and a third amplification target, a synthetic oligonucleotide internal PCR control (IPC), to allow for the assessment of PCR inhibition. We chose the two nuclear targets to provide quantity and fragment-length information relevant to the STR amplification targets commonly used for forensic genotyping. The long target (nuTH01, 170-190 bp) spans the TH01 STR locus and uses a FAM-labeled TaqMan probe for detection. The short nuclear target (nuCSF, 67 bp) is directed at the upstream flanking region of the CSF1PO STR locus and is detected using a VIC-labeled TaqManMGB probe. The IPC target sequence is detected using a NED-labeled TaqManMGB probe. The assay was validated on the Applied Biosystems 7500 Real-Time PCR system, which is optimized for NED detection. We report the results of a developmental validation in which the assay was rigorously tested, in accordance with the current SWGDAM guidelines, for precision, sensitivity, accuracy, reproducibility, species specificity, and stability.     

司法会计应为强化法律监督提供技术保障——暨谈对检察机关司法会计可持续发展

从科学发展观的角度审视,仅满足或停留于审计查账,是阻碍检察机关司法会计发展的主要瓶颈。我国司法会计的发展方向是以鉴定为主导,为诉讼提供有别于审计的涉案会计事实证明,我国诉讼证明制度发展的趋势是对涉案会计事实证明的要求日益严格。检察机关强化法律监督,必然要求涉案会计事实证明能够进一步体现严格的执法精神。检察机关司法会计全面协调可持续发展,更好地为强化法律监督提供技术保障,就必须遵循适应我国司法会计的发展方向、诉讼证明制度的发展趋势和强化法律监督要求等三项主要原则。为此,应当加快以鉴定为主的理论研究和业务规范,建立推进理论研究和业务规范及人才培养的机制。     

Fourier transform infrared analyses of some particulate drug mixtures using a diamond anvil cell with a beam condenser and an infrared microscope.

Although the diamond anvil cell (DAC) has been used in many forensic science laboratories for the analysis of trace evidence, few applications of this technique for the analysis of controlled substances have been reported. This may be due to both an unfamiliarity on the part of forensic drug chemists with this accessory and the nature and quality of spectra that result from use of a DAC on a dispersive instrument. Along with low energy throughput, which results in relatively high noise levels, strong broad diamond absorptions occur. With the use of a Fourier transform infrared instrument, these do not present a problem and nanogram quantities of materials can be analyzed when the DAC is used with an infrared microscope. Since single crystals can be sampled with the DAC, simple physical separations (involving particle-picking) can be used in certain cases to isolate drugs from particulate mixtures for infrared analysis. This method is especially useful for some "difficult" mixtures and residues, and several examples of such analyses involving samples of forensic science interest are presented.     

应用自动化工作站提取常见生物样本DNA

目的建立使用自动化工作站提取法医案件生物样本DNA的方法。方法选用Biomek 3000自动化工作站,采用DNA IQTM系统及Chelex法对法医案件中常见生物样本进行DNA提取,荧光定量技术进行定量,PCR扩增16个STR基因座并与手工提取方法比较。结果与手工提取方法进行比较,选用自动化工作站结合使用DNAIQTM系统及Chelex法提取DNA可获得满意的STR检验结果。结论自动化工作站可用于法医案件中常见生物样本的DNA提取。     

Extraction of human DNA for PCR from chewed residues of betel quid using a novel "PVP/CTAB" method

Residues of chewed betel quid (BQ) are often found on crime scenes in Taiwan and possibly some of the Southeast Asian countries. Although these residues are important biological evidences relating to the suspects, the forensic analysis of BQ evidence has been hindered by failures in extraction of human DNA for PCR analysis. Therefore, it is a prerequisite for relevant forensic casework to establish a reliable method for extracting DNA from chewed BQ residues. Three conventional methods (salt/chloroform, 5% Chelex-100 resin, and QIAamp) were first tested for extraction of human DNA from 33 mock BQ samples, which had been stored for less than two months, and 50 four-year-old forensic BQ samples. PCR amplifications from the HLA-DQA1&PM and the STR loci were then used to test the quality of the extracted DNA. For the mock samples, three observations were made. First, PCR amplification of DNA extracted by using these conventional methods had low success rate. Second, the addition of extra Taq DNA polymerase could compensate the lost enzyme activities due to putative inhibitors and, thus, increase the yield. Third, using the Centricon-100 column to remove putative inhibitors substantially improved the efficiency of PCR. However, for the four-year-old forensic BQ samples, none of the attempts for PCR were successful. In order to solve the problem in PCR analysis of DNA from old BQ samples, we developed a DNA extraction method based on the use of polyvinyl pyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB), which bind to two common classes of PCR inhibitors in plants, polyphenols, and polysaccharides, respectively. The result showed that this "PVP/CTAB" method is completely successful for the mock BQ samples, and 92% (46 out of 50) successful for the four-year-old forensic BQ samples. To our best knowledge, this is the first report of a reliable method for the extraction of human DNA for PCR from chewed BQ residues. This method should provide a useful means for forensic identification in countries where betel chewing is common.