Jeong Sangkyun, Hahn Yoonsoo, Rong Qi, Pfeifer Karl
Laboratory of Mammalian Genes and Development, National Institute of Child Health and Development, National Institutes of Health, Bethesda, Maryland 20892, USA.
Genome Res. 2007 Jul;17(7):1093-100. doi: 10.1101/gr.6028507. Epub 2007 Jun 1.
Epigenetic and genetic mechanisms can result in large differences in expression levels of the two alleles in a diploid organism. Furthermore, these differences may be critical to phenotypic variations among individuals. In this study, we present a novel procedure and algorithm to precisely and accurately quantitate the relative expression of each allele. This method uses the differential melting properties of DNAs differing at even a single base pair. By referring to the melting characteristics of the two pure alleles, the fractional contribution of the two alleles to any unknown mixture can be mathematically resolved. These methods are highly accurate and precise because each single melting reaction yields multiple data points for analysis. Finally, we discuss how this approach can be used more generally to accurately quantitate gene expression relative to known standards.
表观遗传和遗传机制可导致二倍体生物中两个等位基因的表达水平出现巨大差异。此外，这些差异可能对个体间的表型变异至关重要。在本研究中，我们提出了一种新颖的程序和算法，用于精确且准确地定量每个等位基因的相对表达。该方法利用了即使仅相差一个碱基对的DNA的不同解链特性。通过参考两个纯合等位基因的解链特征，可以通过数学方法解析这两个等位基因对任何未知混合物的贡献比例。这些方法高度准确和精确，因为每个单一的解链反应都会产生多个用于分析的数据点。最后，我们讨论了如何更广泛地使用这种方法相对于已知标准来准确地定量基因表达。
