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161.
Dide Boelens Roberta Fogliatto Mariot Mirna Ghemrawi Ate D. Kloosterman Bruce R. McCord 《Electrophoresis》2021,42(12-13):1352-1361
There are situations in which it would be very valuable to have a DNA profile within a short time; for example, in mass disasters or airport security. In previous work, we have promoted reduced size STR amplicons for the analysis of degraded DNA. We also noticed that shorter amplicons are more robust during amplification, making them inhibition resistant, and potentially applicable to high-speed direct PCR. Here, we describe a set of miniSTRs capable of rapid direct PCR amplification. The selected markers are a subset of the Combined DNA Index System (CODIS) loci modified to permit high-speed amplification. Using the proposed protocol, the amplification of eight loci plus amelogenin directly from a saliva sample can be completed in 7 min and 38 s using a two-step PCR with 30 cycles of 98°C for 2 s and 62°C for 7 s on a Streck Philisa thermocycler. Selection of DNA polymerase, optimization of the two-step PCR cycling conditions, the primer concentrations, and the dilution of saliva is described. This method shows great potential as a quick screening method to obtain a presumptive DNA profile when time is limited, particularly when combined with high-speed separation and detection methods. 相似文献
162.
Yiling Qu RuiYang Tao Huan Yu Qi Yang Ziwei Wang Rui Tan Xiaochun Zhang Ruocheng Xia Lei Xiong Shihan Xi Jun Wu Yuzhen Gao Suhua Zhang Chengtao Li 《Electrophoresis》2021,42(14-15):1419-1430
This paper describes the development and validation of a novel 31-locus, six-dye STR multiplex system, which is designed to meet the needs of the rapidly growing Chinese forensic database. This new assay combines 20 extended-CODIS core loci (D3S1358, D5S818, TPOX, CSF1PO, TH01, vWA, D7S820, D21S11, D8S1179, D18S51, D16S539, D13S317, FGA, D1S1656, D2S441, D2S1338, D10S1248, D12S391, D19S433, and D22S1045), nine highly polymorphic loci in Chinese Han population (D3S3045, D6S1043, D6S477, D8S1132, D10S1435, D15S659, D19S253, Penta D, and Penta E), and two gender determining markers, amelogenin and Y-Indel, which could amplify DNA from extracts, as well as direct amplification from substrates. To demonstrate the suitability for forensic applications, this system was validated by precision and accuracy evaluation, concordance tests, case sample tests, sensitivity, species specificity, stability, stutter calculation, and DNA mixtures, according to the guidelines described by the Scientific Working Group on DNA Analysis Methods (SWGDAM) and regulations published by the China Ministry of Public Security. The validation results indicate the robustness and reliability of this new system, and it could be a potentially helpful tool for human identification and paternity testing in the Chinese population, as well as facilitating global forensic DNA data sharing. 相似文献
163.
《Electrophoresis》2017,38(7):1016-1021
A Y‐STR multiplex system has been developed with the purpose of complementing the widely used 17 Y‐STR haplotyping (AmpFlSTR Y Filer® PCR Amplification kit) routinely employed in forensic and population genetic studies. This new multiplex system includes six additional STR loci (DYS576, DYS481, DYS549, DYS533, DYS570, and DYS643) to reach the 23 Y‐STR of the PowerPlex® Y23 System. In addition, this kit includes the DYS456 and DYS385 loci for traceability purposes. Male samples from 625 individuals from ten worldwide populations were genotyped, including three sample sets from populations previously published with the 17 Y‐STR system to expand their current data. Validation studies demonstrated good performance of the panel set in terms of concordance, sensitivity, and stability in the presence of inhibitors and artificially degraded DNA. The results obtained for haplotype diversity and discrimination capacity with this multiplex system were considerably high, providing further evidences of the suitability of this novel Y‐STR system for forensic purposes. Thus, the use of this multiplex for samples previously genotyped with 17 Y‐STRs will be an efficient and low‐cost alternative to complete the set of 23 Y‐STRs and improve allele databases for population and forensic purposes. 相似文献
164.
Quantitative determination of soybean meal content in compound feeds: comparison of near-infrared spectroscopy and real-time PCR 总被引:1,自引:0,他引:1
Standard methods for determining the raw material content of compound feed are little exploited, except for the identification
of meat and bone meal in feeds. In this work, near-infrared (NIR) spectroscopy and real-time polymerase chain reaction (PCR)
were applied in order to establish new and fast methods for quantification of soybean meal content in compound feeds. The
best prediction quality was achieved by using a model based on NIR spectroscopy (R
2 = 0.9857, standard error of cross-validation 1.1065). Furthermore, a sensitive qualitative detection method by using the
real-time PCR was developed (R
2 = 0.976, slope −3.7599). Finally, the differences between the real-time PCR result and the NIR spectroscopy result for a
given sample were also treated, and we found that the NIR spectroscopy method provided quite accurate results which approach
closely those of the real-time PCR method.
Hui Li and Xiaowen Lv contributed equally to this work. 相似文献
165.
A universal hepatitis B virus (HBV) DNA detection kit is appealing for the worldwide diagnosis and monitoring of the treatment
of different mutant types of hepatitis B virus. A sensitive and reproducible real-time PCR assay based on the universal molecular
beacon (U-MB) technique was developed for the detection of HBV DNA in serum. The U-MB probe used in the assay has no interaction
with the HBV DNA sequence. The U-MB technique not only reduced the cost of HBV detection but also had the potential for the
development of a universal detection kit for different mutant HBV types and other DNA systems. To demonstrate its clinical
utility, 90 serum samples were analyzed using the U-MB real-time PCR method. In the experiments we found that several crucial
factors needed to be considered in the primer design, such as the avoidance of formation of severe primer–dimer and primer
self-hairpin structure. With the optimized primer sets, satisfactory results were obtained for all the tested samples. We
concluded that this assay would be an excellent candidate for a universal HBV DNA detection method.
Principle of the U-MB real-time PCR method for HBV DNAdetection 相似文献
166.
Food allergen detection methods and the challenge to protect food-allergic consumers 总被引:1,自引:0,他引:1
van Hengel AJ 《Analytical and bioanalytical chemistry》2007,389(1):111-118
The detection of allergenic ingredients in food products has received increased attention from the food industry and legislative
and regulatory agencies over recent years. This has resulted in the improvement of measures aimed at the protection of food-allergic
consumers. The controlled production of food products and control activities executed by food inspection agencies rely on
the availability of methods capable of detecting traces of allergenic ingredients. The development of such methods faces a
multitude of analytical challenges. Those challenges will be identified and discussed in this review. Furthermore, future
developments and trends in analytical methodology as applied to the detection of food allergens are reported. 相似文献
167.
Chen XW Xu ZR Qu BY Wu YF Zhou J Zhang HD Fang J Wang JH 《Analytical and bioanalytical chemistry》2007,388(1):157-163
Bead injection in a lab-on-valve (LOV) system was adopted for DNA purification via micro solid-phase extraction (SPE) with
a renewable silica microcolumn packed in a channel of the LOV unit. The complex matrix components in human whole blood, including
proteins, were well eliminated by choosing properly the sample loading and elution media. The DNA purification process was
monitored on-line by using laser-induced fluorescence in a demountable side part of the LOV unit incorporating optical fibers.
The practical applicability of the entire system was demonstrated by separation/purification of λ-DNA in a simulated matrix
and human blood genetic DNA by performing SPE, in situ monitoring of the purified products, and postcolumn PCR amplification.
When DNAs in a simulated matrix (10.0 ng μl−1 λ-DNA, 50 ng μl−1 bovine serum albumin, 1.0% Triton X-100) were processed in the present system and laser-induced fluorescence was monitored
at 610 nm, an overall extraction/collection efficiency of 70% was achieved by employing identical sample loading and an elution
flow rate of 0.5 μl s−1, along with a precision of 3.8% relative standard deviation. DNA separation and purification from human whole-blood samples
were performed under similar conditions.
Figure Lab-on-valve mesofluidic system employed for DNA separation and purification integrating a demountable fluorescence flow cell
for in-situ laser induced fluorescence detection 相似文献
168.
A Genosensor for Point Mutation Detection of P53 Gene PCR Product Using Magnetic Particles 下载免费PDF全文
Ezat Hamidi‐Asl Jahan Bakhsh Raoof Mohammad Saeid Hejazi Simin Sharifi Seyed Mahdi Golabi Ilaria Palchetti Marco Mascini 《Electroanalysis》2015,27(6):1378-1386
The development of an electrochemical genosensor involving DNA biotinylated capture probe immobilized on streptavidin coated paramagnetic beads and microfluidic based platform for the detection of P53 gene PCR product is reported. The novelty of this work is the combination of a sensitive electrochemical platform and a proper microfluidic system with a simple and effective enzyme signal amplification technology, ELISA, for detection of target DNA sequence and single nucleotide mutation in p53 tumor suppressor gene sequence. The biosensor has been applied to detect the PCR amplified samples and the results shows that it can discriminate successfully perfect matched DNA from mutant form. 相似文献
169.
Gangyi Chen Wei Wang Yi Yuan Juan Dong Xin Cui Xin Huang Yun Deng Zhuo Tang 《Electrophoresis》2020,41(16-17):1517-1522
Mercury contamination is one of the most serious environmental problems. It can cause serious effects on the human health, such as case damage in the brain, nervous system, immune system, and kidney failure. Therefore, development of an accurate, sensitive, and simple operational detection method for mercury is very necessary. Herein, we report a new strategy for mercury ion detection based on commonly used PCR technique. High selectivity and sensitivity were achieved by the formation of the thymine-Hg-thymine (T-Hg-T) unnatural base pair at the 3’-end of PCR primers. The detection results of PCR amplification in presence of mercury ion could be reported either by using agarose gel analysis or through real-time fluorometric dye tracing for different detection purposes. To our knowledge, this study represents the first application of PCR based technique to the detection of metal ions. 相似文献
170.