Analysis of gene expression in Arabidopsis thaliana by array hybridization with genomic DNA fragments aligned along chromosomal regions.

ABSTRACT

The availability of the entire genomic sequence of the higher plant Arabidopsis thaliana prompted an analysis of chromosomal regions for gene expression with the use of high-density DNA array filters spotted with genomic DNA fragments (genomic DNA array analysis). One TAC and nine P1 clones, each of which contains a genomic DNA insert of approximately 80 kb and was used for sequencing of chromosome 5, were arbitrarily selected for analysis. The total size of the genomic regions corresponding to these clones is 819 kb. A total of 339 DNA fragments (average size, 2.9 kb) that cover contiguously the 10 chromosomal regions was selected and spotted onto nylon filters. The filters were then subjected to hybridization with (33)P-labelled cDNA molecules that had been synthesized from polyadenylated RNA isolated from 3-week-old plants. Quantitative and reproducible measurement of hybridization signals allowed analysis of the transcription of all genes in the targeted regions that were expressed at a level above the limit of detection. The data revealed that the analysed chromosomal regions are rich in active genes, and that they also provided a basis for the identification of novel transcripts whose sequences are not represented in the expressed sequence tag (EST) database.