MICROARRAYS

    To gain a better understanding of changes in gene expression associated with cold acclimation in blueberry, the Rowland laboratory has undertaken a genomics approach based on the analysis of Expressed Sequence Tags (ESTs) and microarrays. Two standard cDNA libraries were constructed previously using RNA from cold acclimated and non-acclimated floral buds of the highbush V. corymbosum cultivar ‘Bluecrop’. Several thousand cDNAs were picked and ~1200 5’-end ESTs were generated from each of the libraries. Based on a contig analysis of the ~2400 ESTs, a set of representative cDNAs have been selected and imprinted onto microarrays to be used to identify cold-responsive genes in various blueberry genotypes. Expression of cold-responsive genes will be compared in cold tolerant and cold sensitive genotypes to determine which of the cold-responsive genes are the best candidates for contributing to cold tolerance.

    Hybridizations of microarrays have been initiated using fluorescent probes prepared from RNA isolated from floral buds of the cold hardy cultivar ‘Bluecrop’. Floral buds for probe preparation have been collected from field plants and cold room-treated greenhouse plants before exposure to low temperatures (non-acclimated) and multiple times after the beginning of cold stress [100, 400, 800, and 1200 hours of chilling (exposure to 0-7ºC) for field plants and 500 and 1000 hours of exposure to 4ºC combined with short photoperiod for cold room plants]. These microarray experiments will be completed followed by hybridizations to fluorescent probes prepared from RNA isolated from floral buds of cold sensitive genotypes, as described above for ‘Bluecrop’. Possible genotypes for these studies include a southern rabbiteye (V. ashei) cultivar ‘Tifblue’, which we have used in the past for many comparisons to ‘Bluecrop’ involving cold tolerance and dehydrin expression, and a southern highbush (V. corymbosum x V. darrowi hybrid) cultivar ‘Gulfcoast’, which we have used in limited comparisons to ‘Bluecrop’ in the past but which is more closely related to ‘Bluecrop’ genetically than is ‘Tifblue’.