Genome annotation by high-throughput 5' RNA end determination.
Complete gene identification and annotation, including alternative transcripts, remains a challenge in understanding genome organization. Such annotation can be achieved by a combination of computational analysis and experimental confirmation. Here, we describe a high-throughput technique, trans-spliced exon coupled RNA end determination (TEC-RED), that identifies 5' ends of expressed genes in nematodes. TEC-RED can distinguish coding regions from regulatory regions and identify genes as well as their alternative transcripts that have different 5' ends. Application of TEC-RED to approximately 10% of the Caenorhabditis elegans genome yielded tags 75% of which experimentally verified predicted 5'-RNA ends and 25% of which provided previously unknown information about 5'-RNA ends, including the identification of 99 previously unknown genes and 32 previously unknown operons. This technique will be applicable in any organisms that have a trans-splicing reaction from spliced leader RNA. We also describe an efficient sequential method for concatenating short sequence tags for any serial analysis of gene expression-like techniques.
Hwang
Byung Joon
BJ
Howard Hughes Medical Institute and Division of Biology, 156-29, California Institute of Technology, 1200 East California Boulevard, Pasadena, CA 91125, USA.
Müller
Hans-Michael
HM
Sternberg
Paul W
PW
D000818:Animals; D001483:Base Sequence; D017173:Caenorhabditis elegans / Q000235:genetics*; D017931:DNA Primers; D016678:Genome; D012313:RNA / Q000235:genetics*
D017931:DNA Primers; D012313:RNA