Detection of allelic imbalance in gene expression using pyrosequencing.
Single-nucleotide polymorphisms (SNPs) are common in the human genome, with more than 11 million SNPs having frequencies greater than 1%. The challenge is to identify the minority of functional SNPs from the large number of SNPs that are expected to be silent. Whereas coding variants are unusual, and functional (nonsynonymous) coding SNPs likely rare, regulatory SNPs appear to be common. Traditional methods to identify these SNPs in vitro are time consuming and challenging. An alternative method is to examine the allele-specific expression in the cDNA from tissues expressing the genes of interest and in individuals heterozygous for a transcribed SNP. This method permits expression to be evaluated in the context of the same trans-acting factors and to identify genes with likely cis-acting regulatory variants or parent of origin (imprinting) effects. Such studies require a method to reliably quantify the expression from each allele. Pyrosequencing offers such capabilities, and given the relatively low cost and high throughput, it offers a sensitive method to determine allelic imbalance in the cDNA from tissues expressing genes of interest.
Wang
Hua
H
Division of Endocrinology, Department of Internal Medicine, Central Arkansas Veterans Healthcare System and College of Medicine, University of Arkansas for Medical Sciences, Little Rock, AR, USA.
Elbein
Steven C
SC
D000483:Alleles; D022981:Allelic Imbalance / Q000235:genetics*; D001483:Base Sequence; D004247:DNA / Q000302:isolation & purification; D017931:DNA Primers / Q000378:metabolism; D018076:DNA, Complementary; D011756:Diphosphates / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics*; D005819:Genetic Markers; D015894:Genome, Human / Q000235:genetics; D005838:Genotype; D006801:Humans; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D012313:RNA / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D048348:Reverse Transcription / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*
D017931:DNA Primers; D018076:DNA, Complementary; D011756:Diphosphates; D005819:Genetic Markers; D012333:RNA, Messenger; D012313:RNA; D004247:DNA