ARS | Publication request: Comparative Analysis of Ests from Cold Acclimated and Non-Acclimated Leaves of Rhododendron.

Submitted to: Meeting Abstract
Publication Acceptance Date: August 1, 2004
Publication Date: August 20, 2004
Citation: Wei, J., Dhanaraj, A., Rowland, L.J., Fu, Y., Krebs, S.L., Arora, R. 2004. Comparative Analysis Of Ests From Cold Acclimated And Non-Acclimated Leaves Of Rhododendron.. Meeting Abstract.

Technical Abstract: An expressed sequence tag (EST) analysis approach was undertaken to characterize the expression of genes involved in cold acclimation of Rhododendron, a broad-leaf, woody evergreen species. Two cDNA libraries were constructed, one from winter-collected (cold acclimated; CA; leaf freezing tolerance '53 ºC) leaves of field-grown Rhododendron catawbiense plants, and the other from non-acclimated (NA; leaf freezing tolerance '7 ºC) leaves collected in the previous summer. cDNA clones were randomly picked from these libraries and single-pass sequenced. The resultant ESTs were evaluated by base-calling and subsequently trimmed of vector and low-quality sequences. A total of 862 5'-end high-quality ESTs were generated from the two libraries, 423 from CA and 439 from NA library, and a contig analysis was performed. About 6.3% of these unique transcripts were shared between the libraries, suggesting marked differences in gene expression between CA and NA leaves. Separate clustering of ESTs from CA and NA libraries was carried out and putative functions were assigned to cDNAs by BLASTX. A total of 674 unique cDNAs (340 from CA and 334 from NA library) were identified, of which 454 had known functions (206 from CA and 248 from NA library). Analysis of the relative frequency at which specific cDNAs were picked from the two libraries indicated that 6 gene families were highly abundant in CA library and included early light-inducible proteins, dehydrins, and beta-amylase among others. Similarly, 8 gene families were highly abundant in NA library including chlorophyll a/b-binding protein, RuBisCO small subunit precursor, RuBisCO activase, and chloroplast precursor of plastocyanin among others. Northern blot analyses for the 4 highly abundant genes selected from each of the cDNA libraries confirmed their preferential expression in either NA or CA leaf tissues. Our results suggest that osmotic regulation, desiccation tolerance, photoinhibition tolerance, and photosynthesis adjustment are some of the key components of cold adaptation in Rhododendron. Several ESTs from the CA library were identified as encoding five distinct dehydrins. Considering that only one major cold-responsive dehydrin had been previously detected in R. catawbiense leaf tissues on immunoblots, EST profiling approach appears to be an effective strategy to identify cold acclimation-responsive transcripts in Rhododendron.