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1.
目的 建立一种能够快速鉴定人参和西洋参的双重实时荧光PCR方法。方法 通过对人参属及其近似种基因序列的分析比对,设计特异性的引物探针,建立双重实时荧光PCR检测方法。PCR反应体系为Premix Ex Taq (Probe qPCR) 10 μL,人参上下游引物各0.5 μL,西洋参上下游引物各0.3 μL,人参与西洋参特异性探针各0.4 μL,DNA模板2 μL,灭菌去离子水补至20 μL。反应条件为95℃预变性30 s;然后以95℃变性5 s,60℃退火延伸30 s进行40个循环。应用该方法测试了27份DNA样品,包括7份人参、6份西洋参、4份人参与西洋参混合样、10份其他人参属样品及其他常见中药材样品的DNA,以确定该方法的特异性。将人参与西洋参样品DNA混合后10倍稀释成不同浓度后进行检测,用以确定该方法的灵敏度。结果 人参及西洋参样品均在特定的荧光通道下出现典型的阳性扩增曲线,人参与西洋参混合样品均同时出现两条阳性扩增曲线,其它对照样品及空白对照均没有出现阳性扩增曲线。灵敏度检测结果显示该方法检测人参及西洋参的最低检测限均为2 pg DNA/反应。结论 本实验建立的双重实时荧光PCR方法能够同时快速、准确、灵敏地鉴别出人参和西洋参。 相似文献
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A developmental validation was performed to demonstrate reliability, reproducibility and robustness of the ANDE System with the FlexPlex assay, including an integrated Expert System, across a number of laboratories and buccal sample variations. Previously, the related DNAscan™/ANDE 4C Rapid DNA System using the PowerPlex®16 assay and integrated Expert System Software received NDIS approval in March 2016. The enhanced ANDE instrument, referred to as ANDE 6C, and the accompanying 6-dye, 27-locus STR assay, referred to as FlexPlex, have been developed to be compatible with all widely used global loci, including the expanded set of the CODIS core 20 loci.Six forensic and research laboratories participated in the FlexPlex Rapid DNA developmental validation experiments, testing a total of 2045 swabs, including those obtained from 1387 unique individuals. The goal of this extensive and comprehensive validation was to thoroughly evaluate and document the ANDE System and its internal Expert System to reliably genotype reference buccal swab samples in a manner compliant with the FBI’s Quality Assurance Standards and the NDIS Operational Procedures.The ANDE System, including automated Expert System analysis, generated reproducible and concordant results for buccal swabs when testing various instruments at different laboratories by a number of different operators. When testing a number of non-human DNAs, including oral bacteria, the ANDE System and FlexPlex assay demonstrated limited cross-reactivity. Potential PCR inhibitors were evaluated as part of the validation and no inhibition was detected. Reproducible and concordant profiles were generated from buccal swab samples collected with a limit of detection appropriate for buccal swab collections from arrestees. The precision and resolution of the System met industry standards for detection of microvariants and single base resolution.The integrated Expert System appropriately demonstrated the ability to correctly pass or fail profiles for CODIS upload without human review. During this comprehensive developmental validation, the ANDE System successfully interpreted over 2000 samples tested with over 99.99% concordant alleles. The data package described herein led to the ANDE System with the FlexPlex assay receiving NDIS approval in June 2018. 相似文献
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目的:建立HPLC/TQ-MS测定益母草中阿魏酸含量的方法并测定不同产地益母草中阿魏酸含量,评价益母草中阿魏酸对中成药调经止痛片及新生化颗粒的质量影响,为制定含有益母草并以阿魏酸为定量指标的中成药质量标准提供参考。方法:采用薄层色谱法及UPLC-Q/TOF定性鉴别及分析益母草中阿魏酸,采用HPLC/TQ-MS定量分析10批不同产地益母草中阿魏酸含量。基于定量分析结果及处方用量探讨益母草中阿魏酸含量对其中成药质量标准的影响。结果:薄层鉴别及UPLC-Q/TOF定性分析结果显示,10批不同产地益母草中均含有阿魏酸。HPLC/TQ-MS定量分析结果显示,益母草中阿魏酸含量在0.004 2%~0.006 5%之间。结论:益母草与当归、川芎等含有阿魏酸的药材配伍且其用量较大时,益母草中阿魏酸含量会对其质量评价产生一定的影响。 相似文献
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Nathan Scudder James Robertson Sally F. Kelty Simon J. Walsh Dennis McNevin 《The Australian journal of forensic sciences》2020,52(2):235-241
ABSTRACTForensic DNA analysis is dependent on comparing the known and the unknown. Expand the number of known profiles, and the likelihood of a successful match increases. Forensic use of DNA is moving towards comparing samples of unknown origin with publicly available genetic data, such as the records held by genetic genealogy providers. Use of forensic genetic genealogy has yielded a number of recent high-profile successes but has raised ethical and privacy concerns. Navigating family trees is complex, even more so when combined with a comparison of genetic relationships. This intelligence-gathering process has led to occasional false leads, and its use also risks a public backlash, similar to concerns over Cambridge Analytica. A cautious approach to use of this technique is therefore warranted. 相似文献
7.
By substituting the saturated vapour phase tension of the pure normal hydrocarbons described by the Clausius-Clapeyron law into the accepted expression of the specific retention volume (Vg,T), a theoretically coherent and relatively simple mathematical evidence of the elution behaviour of the homologous members has been deduced. It gives exponential retention time dependence on carbon number for isothermal, and nearly equidistant (i.e., approximately linear retention-time dependence on carbon number) elution for linear temperature programmed gas chromatographic runs. The final equations are in close correlation with the experimental results. Special emphasis is placed on the fact that a good approximation—not strict physical laws—have been found. 相似文献
8.
Â. C. Volpini M. J. Marques S. Lopes dos Santos G. L. Machado-Coelho W. Mayrink A. J. Romanha 《Clinical microbiology and infection》2006,12(8):815-818
This study examined the ability of PCR to amplify Leishmania DNA, stored on Giemsa-stained slides, from American cutaneous leishmaniasis (ACL) patients. In total, 475 slides stored for up to 36 years were obtained from an outpatient clinic in a Brazilian ACL-endemic region, and Leishmania DNA was amplified from 395 (83.2%) of the DNA samples using primers specific for the minicircle kinetoplast DNA. Restriction fragment length polymorphism analysis of these amplicons demonstrated that Leishmania (Viannia) braziliensis was the only species present in these samples. The results demonstrated that archived Giemsa-stained slides can provide a Leishmania DNA source for performing clinical and epidemiological studies of leishmaniasis. 相似文献
9.
A. G. G. Lødeng C. Ahlén H. Lysvand L. H. Mandal O. J. Iversen 《Clinical microbiology and infection》2006,12(8):761-768
This report describes a new PCR-based assay for the detection of Pseudomonas aeruginosa genotype D in occupational saturation diving systems in the North Sea. This genotype has persisted in these systems for 11 years (1993-2003) and represents 18% of isolates from infections analysed during this period. The new PCR assay was based on sequences obtained after randomly amplified polymorphic DNA (RAPD)-PCR analysis of a group of isolates related to diving that had been identified previously by pulsed-field gel electrophoresis (PFGE). The primer set for the D genotype targets a gene that codes for a hypothetical class 4 protein in the P. aeruginosa PAO1 genome. A primer set able to detect P. aeruginosa at the species level was also designed, based on the 23S-5S rDNA spacer region. The two assays produced 382-bp and 192-bp amplicons, respectively. The PCR assay was evaluated by analysing 100 P. aeruginosa isolates related to diving, representing 28 PFGE genotypes, and 38 clinical and community P. aeruginosa isolates and strains from other species. The assay identified all of the genotype D isolates tested. Two additional diving-relevant genotypes (TP2 and TP27) were also identified, as well as three isolates of non-diving origin. It was concluded that the new PCR assay is a useful tool for early detection and prevention of infections with the D genotype. 相似文献
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短串联重复序列HumFGA、D3S1359的法医学应用研究 总被引:1,自引:0,他引:1
对短串联重复序列(short tandem repeats,STR)的两个高多态位点HumFGA(human alpha fibrinogen,人类α-纤维蛋白原基因)、D3S1359的法医学应用进行研究,并通过实际检测案件统计结果进行评估。实验结果表明:两位点灵敏度高,分别为0.2、0.5ngDNA;同一性和可重复性均较理想;种属特异性好,常见动物未发现扩增产物,仅灵长类动物(黑叶猴和猕猴)有扩增条带;复合扩增体系效果良好;在352次减数分裂中,D3S1359未发现突变,HumFGA检测到1次,突变率为0.28%;实际案件统计和应用结果也显示两位点是多态性高,实用性强的两个遗传标记系统。HumFGA和D3S1359在法医学个人识别与亲子鉴定案例中有较大应用价值。 相似文献