Rapid genotyping of factor V Leiden mutation using single-tube bidirectional allele-specific amplification and automated ultrathin-layer agarose gel electrophoresis

Sasvári-Székely, Mária; Gerstner, Árpád; Rónai, Zsolt; Staub, Maria; Guttman, András

doi:10.1002/(sici)1522-2683(20000301)21:4<816::aid-elps816>3.0.co;2-y

Cited by 22 publications

(10 citation statements)

References 25 publications

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“…DNA samples were taken from healthy Hungarian volunteers using noninvasive DNA sampling (buccal swabs) and conventional phenol extraction of DNA was used, as previously described [25,26]. The Qiagen HotStar Taq DNA polymerase kit was used for PCR amplification.…”

Section: Dna Isolation and Pcr Amplificationmentioning

confidence: 99%

“…Allele-specific amplification is one of the most efficient methods of SNP genotyping [3].This technique utilizes the characteristics of the DNA polymerases, which can extend the 3'-end of the primer only when there is a perfect match to the template DNA, provided that the enzyme does not possess 3'?5' exonuclease activity. If the SNPs of interest have two forms in the human population, two allele-specific primers should be used, either in two separate PCR reactions [4] or in their combination [5,6].…”

Section: Introductionmentioning

confidence: 99%

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Direct haplotype detection of adjacent polymorphic sites in the regulatory region of the dopamine D4 receptor (DRD4) gene

et al. 2002

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A novel method of haplotype identification is discussed allowing simultaneous detection of two adjacent polymorphic sites, a single nucleotide polymorphism (SNP) and a length polymorphism within 1-2 kilobase distance. The method combines allele specific-amplification with high-throughput, automated ultrathin-layer gel electrophoresis analysis of fragment size polymorphism. A typical application is shown for genotyping the -521 C/T single nucleotide polymorphism and the 120 bp duplication in the 5"-upstream region of the dopamine D4 receptor (DRD4) gene. We have also demonstrated that the haplotypes of double heterozygotes for -521 C/T and for the 120 bp duplication can be clearly distinguished, that has only been possible previously by extensive pedigree analysis.

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Section: Dna Isolation and Pcr Amplificationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Direct haplotype detection of adjacent polymorphic sites in the regulatory region of the dopamine D4 receptor (DRD4) gene

et al. 2002

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“…DNA samples were taken from healthy Hungarian volunteers using noninvasive DNA sampling and conventional phenol extraction of DNA, as previously described [17,18]. The Qiagen HotStar Taq DNA polymerase kit was used for PCR amplification of the region around the 5-HTTLPR.…”

Section: Dna Source and Pcr Amplificationmentioning

confidence: 99%

High‐throughput genotyping of repeat polymorphism in the regulatory region of serotonin transporter gene by gel microchip electrophoresis

Nemoda¹,

Rónai²,

Székely³

et al. 2001

Electrophoresis

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Large-scale genotyping of the repeat polymorphism in the regulatory region of the serotonin transporter gene (5-HTTLPR) was attempted by polymerase chain reaction (PCR) amplification followed by gel microchip electrophoresis analysis. The multilane (96) format of the gel microchip system allowed parallel separation of a large number of samples. The separation and visualization of the PCR amplicons from either the 5-HTTLPR short allele (number of repeats are 14) or the 5-HTTLPR long form (16 repeats) was completed in a few minutes. Genotyping of healthy Caucasian individuals showed that the short allele had a somewhat lower frequency (0.42) than the long form (0.58), and the genotype frequencies fulfilled the criteria of the Hardy-Weinberg equilibrium (chi = 0.012, p = 0.994). Based on these results, gel microchip electrophoresis system proved to be a powerful tool for high throughput genotyping of repeat polymorphism.

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“…The two outer primers amplify a constant fragment, which is also a PCR positive control. Once again, this method required optimization because of the effects of competition among multiple primer sets in the PCR reaction (Liu et al 1997;Sasvari-Szekely et al 2000;Waterfall and Cobb 2001;Waterfall and Cobb 2002). Moreover, the efficient amplification of all expected fragments was not always guaranteed (Waterfall and Cobb 2001).…”

Section: Introductionmentioning

confidence: 99%

Single-reaction for SNP Genotyping on Agarose Gel by Allele-specific PCR in Black Poplar (Populus nigra L.)

et al. 2007

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The wide development of single nucleotide polymorphism (SNP) markers also in non-model species increases the need for inexpensive methods that do not require sophisticated equipment and time for optimization. This work presents a new method for polymerase chain reaction (PCR) amplification of multiple specific alleles (PAMSA), which allows efficient discrimination of SNP polymorphisms in one reaction tube with standard PCR conditions. This improved PAMSA requires only three unlabeled primers: a common reverse primer and two allele-specific primers having a tail of different length to differentiate the two SNP alleles by the size of amplification products on agarose gel. A destabilizing mismatch within the five bases of the 3′ end is also added to improve the allele specificity. To validate the accuracy of this method, 94 full-sib individuals were genotyped with three SNPs and compared to the genotypes obtained by cleaved amplified polymorphic sequence (CAPS) or derived CAPS. This method is flexible, inexpensive, and well suited for high throughput and automated genotyping.

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Rapid genotyping of factor V Leiden mutation using single-tube bidirectional allele-specific amplification and automated ultrathin-layer agarose gel electrophoresis

Cited by 22 publications

References 25 publications

Direct haplotype detection of adjacent polymorphic sites in the regulatory region of the dopamine D4 receptor (DRD4) gene

Direct haplotype detection of adjacent polymorphic sites in the regulatory region of the dopamine D4 receptor (DRD4) gene

High‐throughput genotyping of repeat polymorphism in the regulatory region of serotonin transporter gene by gel microchip electrophoresis

Single-reaction for SNP Genotyping on Agarose Gel by Allele-specific PCR in Black Poplar (Populus nigra L.)

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