High-throughput detection of multiple genetic polymorphisms influencing drug metabolism with mismatch primers in allele-specific polymerase chain reaction

Ishiguro, Akihiro; Kubota, Takeshi; Soya, Yoshihiro; Sasaki, Hiroshi; Yagyu, Osamu; Takarada, Yutaka; Iga, Tatsuji

doi:10.1016/j.ab.2004.11.038

Cited by 41 publications

(23 citation statements)

References 27 publications

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“…The genotyping of the SNP (808G>T) of SLC22A2 was performed using the ASP-PCR method according to previous reports [18] . The primer set (forward primer: 5′-AAAGGTTCTACCGTCCAA-3′; reverse primer for 808G: 5′-GTGGTTGCAGTTCACAGTAGC-3′; reverse primer for 808T: 5′-GGTGGTTCCAGTTCACAG-TATC-3′) was designed to amplify a 359-bp and 360-bp portion of the SLC22A2 gene encoding the SNP.…”

Section: Snp Determination (808g>t) In the Slc22a2 Gene Using The Aspmentioning

confidence: 99%

“…The allele-specific primer polymerase chain reaction (ASP-PCR) assay is a method used to determine SNPs based on DNA amplification, and the SNPs can be determined by whether the primers anneal to the target DNA [17,18] . The present study was designed to determine SNPs at position 808 of the SLC22A2 gene using ASP-PCR and to investigate the potential relationship between SLC22A2 gene polymorphisms and blood lactate concentrations in Shanghai Hans with T2DM.…”

Section: Introductionmentioning

confidence: 99%

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SLC22A2 gene 808 G/T variant is related to plasma lactate concentration in Chinese type 2 diabetics treated with metformin

Liu

Zheng

et al. 2010

Acta Pharmacol Sin

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Aim: To investigate the potential relationship between the SLC22A2 gene polymorphism and blood lactate concentration in Shanghai Hans suffering from type 2 diabetes mellitus (T2DM). Methods: The SLC22A2 single nucleotide polymorphism (SNP) 808G/T was genotyped in 400 T2DM patients, including a metformintreated group (n=200) and a non-metformin-treated group (n=200). Fasting plasma lactic acid levels were measured with an enzymeelectrode assay. Biochemical indexes, including plasma alanine aminotransferase (ALT), creatinine (Cr), and glycolated hemoglobin (HbA1c), were also measured. Results: The fasting plasma lactate concentration in the metformin-treated group was significantly higher than that in the non-metformin-treated group (1.29±0.45 mmol/L vs 1.18±0.44 mmol/L, P=0.015). Additionally, the ratio of patients with hyperlactacidemia was 8% (16/200) for the metformin-treated group and 5.5% (11/200) for the non-metformin-treated group, with no lactic acidosis found in either group. The frequency of the SLC22A2 808G/T T allele was 12.9%. Patients with the mutant genotype (TT) had a higher blood lactate concentration in the metformin-treated group than those in the non-metformin-treated group (t=2.492, P=0.013). This trend was not observed in the GG and GT genotypes when compared with metformin-treated and non-metformin-treated groups. Patients with the mutant genotype (TT) in the metformin-treated group also had a higher incidence of hyperlactacidemia compared with the GG genotype (40.0% vs 6.9%, P=0.050) in the metformin-treated group and the GG (6.0%, P=0.042) or GT (4.3%, P=0.043) genotypes in the non-metformin-treated group. In the metformin-treated group, there were significant gender differences in lactate concentrations in the TT (2.18±0.15 vs 1.04±0.27 mmol/L, P=0.008) and GG genotypes (1.40±0.51 vs 1.19±0.35 mmol/L, P=0.004). The lactate levels of women with the TT genotype were the highest in the metformin-treated group, but differences in lactate levels among the genotypes were not observed in the non-metformin-treated group. Conclusion: There is an 808G/T polymorphism in the SLC22A2 gene in Chinese Hans with T2DM. The 808G>T variance in the SLC22A2 gene can affect the plasma lactate level and the incidence of hyperlactacidemia in T2DM patients undergoing metformin therapy. Additionally, the female patients carrying the TT genotype are prone to lactatemia.

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Section: Snp Determination (808g>t) In the Slc22a2 Gene Using The Aspmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

SLC22A2 gene 808 G/T variant is related to plasma lactate concentration in Chinese type 2 diabetics treated with metformin

Liu

Zheng

et al. 2010

Acta Pharmacol Sin

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“…For example, CYP2D6*4, *5, and *14 lack enzymic activity, while CYP2D6*10 and *21 show decreased enzymic activity (14)(15)(16). Approximately 7% of Caucasians are classified as PM and considered to have CYP2D6*4 or *5, while the frequency of PM in the Japanese population is under 1%, and the main mutations are CYP2D6*5 and *14 (17)(18)(19)(20)(21)(22)(23). In contrast, the frequency of IM is high in Japanese at 15%, which can be explained by CYP2D6*10 (24,25).…”

Section: Introductionmentioning

confidence: 99%

Digital PCR for determination of cytochrome P450 2D6 and sulfotransferase 1A1 gene copy number variations

Motoi

Watanabe²,

Honma

et al. 2017

DD&T

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“…The SNP site at the 1075th position is adenine for the wild-type allele (A allele) and cytosine for the mutant allele (C allele). According to the allele-specific PCR reported for the 2C9*3 alleles, [6] two HP-3-labeled forward primers, FW-1075T and FW-1075G, which had thymine and guanine in the second position from the 3' end, respectively, were used for the PCR (Figure 3 A). To increase the allele specificity, an additional T-T mismatch was introduced at the third position.…”

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confidence: 99%

Secondary‐Structure‐Inducible Ligand Fluorescence Coupled with PCR

et al. 2009

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Single-stranded DNA can change its structure dynamically in response to the presence of a complementary strand. The large structural change from the hairpin secondary structure to the double-stranded form leads to the chemistry of molecular beacons (MBs), or hairpin DNA oligomers having both a fluorescent and a quenching chromophore in the hairpin stem.[1] MBs can be used to report the presence of complementary strands by measuring the increasing fluorescence because of the decreasing energy transfer efficiency. Most DNA probes for fluorescent detection are covalently bound to fluorophores to make energy transfer effective. [1, 2] We have focused on an alternative way to link the fluorescence change with the structural changes of DNA that are induced during polymerase chain reaction (PCR) amplification.[3] We describe herein the chemical concept of noncovalent fluorescent DNA labeling by ligand binding to the secondary structure, which allows us to monitor PCR progress by measuring the change in fluorescence emitted from ligand-primer complexes in a homogeneous solution. This concept is not only complementary to that using fluorescent dye bound selectively to PCR product duplexes, [4] but it also expands the possibility of real-time PCR.The concept of DNA labeling by secondary-structureinducible ligand fluorescence is shown in Figure 1. PCR primers are labeled with a hairpin tag containing cytosine bulges (C-bulges). The molecule 2,7-diamino-1,8-naphthyridine (DANP) binds to a C-bulge in its protonated form (DANPH + ) and emits fluorescence at 430 nm with a 30 nm bathochromic shift from the fluorescence of free, unbound DANP.[5] We hypothesized that, as the PCR progresses, the hairpin tag will dissolve and be transformed into a duplex, thus resulting in the loss of the DANP binding site and a decrease in the fluorescence at 430 nm. Toward this end, we investigated the hairpin tags. These tags should identify the DANP binding site without disturbing the fluorescence efficiency, should not interfere with the PCR, should be transformed effectively into the duplex during PCR, and should be applicable to diverse primers. First, the sequence flanking the C-bulge producing the highest fluorescence intensity was investigated by measuring DANP fluorescence with duplexes having a C-bulge flanked by A-T and T-A base pairs. The G-C and C-G base pairs were omitted from the flanking base pairs because of their quenching of DANP fluorescence.[5b] The characteristic spectrum with a broad single peak at 430 nm was obtained for the A_A/TCT sequence. The relative fluorescence intensity at 450 nm for the four C-bulge duplexes and a fully matched duplex are shown in Figure S1 in the Supporting Information. The C-bulge in the A_A/TCT sequence enhanced DANP fluorescence by 7.6-fold compared with the fully matched duplex and by 82-fold compared with free, unbound DANP.We then designed hairpin tags comprising one to three A_A/TCT units separated by three to five base pairs and a T4 hairpin loop (Table 1). The intensity of DANP fluores...

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High-throughput detection of multiple genetic polymorphisms influencing drug metabolism with mismatch primers in allele-specific polymerase chain reaction

Cited by 41 publications

References 27 publications

SLC22A2 gene 808 G/T variant is related to plasma lactate concentration in Chinese type 2 diabetics treated with metformin

SLC22A2 gene 808 G/T variant is related to plasma lactate concentration in Chinese type 2 diabetics treated with metformin

Digital PCR for determination of cytochrome P450 2D6 and sulfotransferase 1A1 gene copy number variations

Secondary‐Structure‐Inducible Ligand Fluorescence Coupled with PCR

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