The most important product of the sequencing of a genome is a complete, accurate catalogue of genes and their products, primarily messenger RNA transcripts and their cognate proteins. Such a catalogue cannot be constructed by computational annotation alone; it requires experimental validation on a genome scale. Using 'exon' and 'tiling' arrays fabricated by ink-jet oligonucleotide synthesis, we devised an experimental approach to validate and refine computational gene predictions and define full-length transcripts on the basis of co-regulated expression of their exons. These methods can provide more accurate gene numbers and allow the detection of mRNA splice variants and identification of the tissue- and disease-specific conditions under which genes are expressed. We apply our technique to chromosome 22q under 69 experimental condition pairs, and to the entire human genome under two experimental conditions. We discuss implications for more comprehensive, consistent and reliable genome annotation, more efficient, full-length complementary DNA cloning strategies and application to complex diseases.
The nucleotide sequences involved in the illegitimate recombination of four recombinants between bacteriophage lambda DNA and pBR322 in E. coli (lambda TA6, lambda KA3, lambda TA1R, and lambda KA7) were determined. Each resulted from recombination between regions of homology of 10 to 13 base pairs. The presence of a recA+ allele was found to stimulate recombination between lambda DNA and pBR322 approximately 10-fold. Lambda TA6, lambda KA3, and lambda KA7 were isolated in the presence of a recA+ allele and therefore, may have been generated by the recA recombination system. However, lambda TA1R was isolated in a recA mutant, and was presumably generated by a different recombination system. The possibility that it was generated by DNA gyrase is discussed. Two recombination events were required to form lambda KA7, which may indicate that it also was generated by DNA gyrase.
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