High-throughput chemiluminometric genotyping of single nucleotide polymorphisms of histamine, serotonin, and adrenergic receptor genes

被引:6
作者
Toubanaki, Dimitra K. [1 ]
Christopoulos, Theodore K. [1 ,2 ]
Ioannou, Penelope C. [3 ]
Flordellis, Christodoulos S. [4 ]
机构
[1] Univ Patras, Dept Chem, Patras 26500, Greece
[2] Inst Chem Engn & High Temp Chem Proc, Fdn Res & Technol Hellas, Patras 26504, Greece
[3] Univ Athens, Dept Chem, Athens 15771, Greece
[4] Univ Patras, Sch Med, Dept Pharmacol, Patras 26504, Greece
关键词
Single nucleotide polymorphisms; Histamine receptor; Serotonin receptor; Adrenergic receptor; Chemiluminescence; OLIGONUCLEOTIDE LIGATION ASSAY; BETA(3)-ADRENERGIC-RECEPTOR GENE; TRP64ARG POLYMORPHISM; CLOZAPINE RESPONSE; DRUG-THERAPY; ASSOCIATION; SCHIZOPHRENIA; VARIANTS; 5-HT2A; ONSET;
D O I
10.1016/j.ab.2008.10.032
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Several pharmacogenetic studies are focused on the investigation of the relation between the efficacy of various antipsychotic agents (e.g., clozapine) and the genetic profile of the patient with ail emphasis on genes that code for neurotransmitter receptors such as histamine, serotonin, anti adrenergic receptors. We report a high-throughput method for genotyping of single nucleotide polymorphisms (SNPs) within the genes of histamine H2 receptor (HRH2), serotonin receptor (HTR2A1 and HTR2A2), and beta(3) adrenergic receptor (ADRB3). The method combines the high specificity of allele discrimination by oligonucleotide ligation reaction (OLR) and the superior sensitivity anti simplicity of chemiluminometric detection in a microtiter well assay configuration. The genomic region that spans the locus of interest is first amplified by polymerase chain reaction (PCR). Subsequently, an oligonucleotide ligation reaction is performed using a biotinylated common probe anti two allele-specific probes that are labeled at the 3' end with digoxigenin anti fluorescein. The ligation products are immobilized in polystyrene wells via biotin-streptavidin interaction, and the hybrids are denatured. Detection is accomplished by the addition of alkaline phosphatase-conjugated anti-digoxigenin or anti-fltlorescein antibodies in combination with a chemiluminogenic substrate. The ratio of the luminescence signals obtained from digoxigenin anti fluorescein indicates the genotype of the sample. The method was applied Successfully to the genotyping of 23 blood samples for all four SNPs. The results were in concordance with both PCR-restriction fragment length polymorphism analysis anti sequencing. (C) 2008 Elsevier Inc. All rights reserved.
引用
收藏
页码:34 / 41
页数:8
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