Plant & Animal Genome IX Conference Plant & Animal Genome IX Conference
Poster: Sequencing & EST
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A full-length (FL) cDNA library construction and sequencing strategy was used to find genes and to build an integrated functional genomics platform for Arabidopsis and other plant species. An analysis of the population of FL cDNAs revealed the nature of the UTRs, codon usage and encoded protein products. Nucleic acid statistics from the population of FL cDNAs were used to design algorithms to find and predict the structure of genes, in the genomic DNA, for which FL cDNAs were not sequenced. PCR products made from the FL cDNAs and the other identified genes were used to build microarrays and to identify genes represented by restriction fragments on comprehensive cDNA-AFLP gels. Analyses of many mRNA preparations using these microarrays and gels have enabled relative mRNA transcript levels to be determined for most of the genes in plant organs grown under various defined conditions. Gene-specific primers were generated from the cDNAs as well as the predicted genes and used in our highly automated Reverse Genetics program to identify knock-out mutants in a comprehensive collection of T-DNA inserts. The FL cDNAs provided clones for a large fraction of the Arabidopsis ORFs and these are readily added to expression vectors to study the effects of expressing each ORF in specific cells, tissues, organs and environments. This integrated program led to the design and implementation of similar high-throughput strategies for FL cDNA and protein sequence determinations in other plant species for which the genomic sequences are not available. Specific examples of discoveries about Arabidopsis genes and transcripts will be given and the efficiencies of the integrated platform based on cDNAs will be described.