A strategy for the identification of candidate genes for alcohol-related phenotypes and other human disorders using rapid polymerase chain reaction mapping of gene-based sequence-tagged sites.
We describe a method for the rapid identification and mapping of human genes, including those possibly contributing to disease and alcohol-related phenotypes. New human genes are identified from cDNA libraries through single-pass sequencing into the 3' untranslated (3'UT) regions of human brain cDNAs. Primers derived from the 3'UT region sequences [representing gene-based, sequence-tagged sites (STSs)] are used for polymerase chain reaction (PCR) analyses of the CEPH megabase insert yeast artificial chromosome (YAC) DNA pools. With this approach, approximately 18,000 megabase YACs can be screened and a single YAC identified using only 52 PCR reactions. The YAC localization in conjunction with other mapping techniques, such as PCR mapping to human chromosomes using somatic cell hybrids, allows identification of chromosomal band locations. In this manner, each gene can be associated with its own STS, which in turn specifies both a corresponding genomic clone and specific location in the genome. These locations can be compared with the purported locations of disease genes. The locations of the STSs can also be compared with those of Quantitative Trait Loci implicated for quantitative traits (e.g., alcohol-related phenotypes) on the basis of synteny between the mouse and human genes. Using this strategy, we found candidates for 78 human disease/syndrome genes among the first 220 genes mapped.
D000437:Alcoholism / Q000235:genetics*; D000818:Animals; D001921:Brain / Q000473:pathology; D002874:Chromosome Mapping; D018244:Chromosomes, Artificial, Yeast; D018076:DNA, Complementary / Q000235:genetics; D015723:Gene Library; D005819:Genetic Markers / Q000235:genetics*; D006801:Humans; D051379:Mice; D008957:Models, Genetic; D010641:Phenotype; D016133:Polymerase Chain Reaction / Q000379:methods*; D016324:Sequence Tagged Sites
D018076:DNA, Complementary; D005819:Genetic Markers