CLONING AND CHARACTERISATION OF THE CDNA CLONES OF THREE GENES THAT ARE DIFFERENTIALLY EXPRESSED DURING DORMANCY-BREAKAGE IN THE SEEDS OF DOUGLAS FIR (PSEUDOTSUGA MENZIESII)

摘要

Subtractive hybridisation methodology and differential screening have been used to isolate cDNA clones of genes that show enhanced expression during cold, moist dormancy-breakage treatment (stratification) of Douglas fir (Pseudotsuga menziesii) seeds. This paper describes the isolation and characterisation of three of these cDNA clones. Sequence analysis reveals a high degree of similarity between the three cDNA clones and Late Embryogenesis Abundant (LEA) protein genes. Each of the three Douglas fir clones encodes a different class of LEA protein (Dure et al., 1989). One of the genes (DF65), showing strong sequence similarity to a dehydrin (class 2 LEA) gene, is expressed in seeds treated at 4 degrees C but not in those seeds treated at 20 degrees C. The other two genes (DF6 and DF77), which are highly similar to class 1 and class 3 LEA genes respectively, were expressed in a similar pattern during dormancy-breakage, with expression at 4 degrees C being maintained at a high level for four weeks of treatment, whereas in seeds incubated at 20 degrees C, expression level decreased after one week of treatment. It is shown that there is a family of genes related to DF65 and DF77 expressed in Douglas fir seeds. Possible roles for LEA proteins in dormancy-breakage of seeds is discussed.