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Genotyping of singl...
Genotyping of single nucleotide polymorphisms using base-quenched probe: A method does not invariably depend on the deoxyguanosine nucleotide
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Luo, Guanghua (författare)
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Zheng, Lu (författare)
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Zhang, Xiaoying (författare)
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Zhang, Jun (författare)
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- Nilsson-Ehle, Peter (författare)
- Lund University,Lunds universitet,Avdelningen för klinisk kemi och farmakologi,Institutionen för laboratoriemedicin,Medicinska fakulteten,Division of Clinical Chemistry and Pharmacology,Department of Laboratory Medicine,Faculty of Medicine
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- Xu, Ning (författare)
- Lund University,Lunds universitet,Avdelningen för klinisk kemi och farmakologi,Institutionen för laboratoriemedicin,Medicinska fakulteten,Division of Clinical Chemistry and Pharmacology,Department of Laboratory Medicine,Faculty of Medicine
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(creator_code:org_t)
- Elsevier BV, 2009
- 2009
- Engelska.
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 1096-0309 .- 0003-2697. ; 386:2, s. 161-166
- Relaterad länk:
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http://dx.doi.org/10...
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https://lup.lub.lu.s...
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https://doi.org/10.1...
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Abstract
Ämnesord
Stäng
- Most available methods for detecting single nucleotide polymorphisms (SNPs) are based principally on the system that can produce an increased fluorescence signal during hybridization. In the current study, we demonstrate a method of base-quenched probe for polymerase chain reaction (PCR) genotyping that requires only a pair of primers and one fluorescent probe and does not invariably depend on the deoxyguanosine nucleotide. This method further exploits the phenomenon of fluorescence quenching of fluorescent-labeled probe during hybridization to its complementary target gene's sequence. 6-Carboxyfluorescein (FAM) can be directly conjugated to a base of either adenine (A), thymine (T), cytosine (C), or guanine (G), referred to as A-, T-, C-, or G-quenched probe, respectively, at either the 5' or 3' end. For describing the method in detail, we chose apolipoprotein M (apoM) as a target gene in the current study. DNA sequencing analyses validated that all four types of base-quenched probes could provide unbiased genotyping results (K = 1, P = 0.000), although the maximum speed of fluorescence increase, max(dF/dT), when using the G-quenched probe method, was approximately twofold lower than the others (P < 0.0001). Moreover, we applied this method to detect another seven SNPs in the genomes of phospholipase A2, monocyte chemoattractant protein I (MCP1), and L-ficolin, further confirming our method. It is concluded that this method is precise, simple, and economic as well as suitable for large-scale genotyping Studies. (C) 2008 Elsevier Inc. All rights reserved.
Ämnesord
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinska och farmaceutiska grundvetenskaper -- Farmakologi och toxikologi (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Basic Medicine -- Pharmacology and Toxicology (hsv//eng)
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinska och farmaceutiska grundvetenskaper -- Läkemedelskemi (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Basic Medicine -- Medicinal Chemistry (hsv//eng)
Nyckelord
- Polymerase chain reaction
- Fluorescence quenching
- Single nucleotide
- polymorphism
Publikations- och innehållstyp
- art (ämneskategori)
- ref (ämneskategori)
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