Täpp I, Malmberg L, Rennel E, Wik M, Syvänen A C
Uppsala University, Sweden.
Biotechniques. 2000 Apr;28(4):732-8. doi: 10.2144/00284rr02.
Homogeneous assays based on real-time fluorescence monitoring during PCR are relevant alternatives for large-scale genotyping of single-nucleotide polymorphisms (SNPs). We compared the performance of the homogeneous TaqMan 5'-nuclease assay and the Molecular Beacon assay using three SNPs in the human estrogen receptor gene as targets. When analyzing a panel of 90 DNA samples, both assays yielded a comparable power of discrimination between the genotypes of a C-to-T transition in codon 10 and a G-to-A transition in codon 594 of the estrogen receptor gene. The Molecular Beacon probes distinguished better than the TaqMan probes between homozygous and heterozygous genotypes of a C-to-G transversion in codon 325. The sensitivity of detecting one allele, present as a minority in a mixed sample, varied between the SNPs and was similar for both assays. With the Molecular Beacon assay, the measured signal ratios were proportional to the amount of the minor allele over a wider range than with the TaqMan assay at all three SNPs.
基于PCR过程中实时荧光监测的均相分析方法是单核苷酸多态性（SNP）大规模基因分型的相关替代方法。我们以人类雌激素受体基因中的三个SNP为靶点，比较了均相TaqMan 5'-核酸酶分析和分子信标分析的性能。在分析一组90个DNA样本时，两种分析方法在雌激素受体基因第10密码子的C到T转换和第594密码子的G到A转换的基因型之间产生了相当的区分能力。在第325密码子的C到G颠换的纯合子和杂合子基因型之间，分子信标探针比TaqMan探针区分得更好。检测混合样本中作为少数存在的一个等位基因的灵敏度在不同SNP之间有所不同，两种分析方法相似。对于分子信标分析，在所有三个SNP处，测量的信号比在比TaqMan分析更宽的范围内与次要等位基因的量成比例。
