Cloning, characterization and expression of dormancy-associated genes in Avena fatua

Bailin Li, Purdue University

Abstract

Dormant seeds of wild oat (Avena fatua) line M73 do not germinate when imbibed in water even when conditions are favorable for germination. Afterripening (dry warm conditions) breaks seed dormancy. To test the hypothesis that differential gene expression is responsible for the maintenance and/or termination of seed dormancy, we compared the polypeptide patterns of dormant and afterripened embryos. Polypeptides labeled in vivo or translated in vitro were separated by two-dimensional electrophoresis followed by fluorography. Many of the newly synthesized proteins were more abundant in dormant embryos, while only a few were more prevalent in afterripened embryos. Furthermore, we isolated cDNA clones corresponding to genes that were differentially expressed in dormant and afterripened embryos. Gene transcripts of these clones were maintained in embryos of imbibed dormant caryopses, but declined rapidly in afterripened embryos after imbibition. GA$\sb3$ treatment of dormant caryopses, which breaks dormancy, could lower the transcript levels in dormant embryos. When the germination of afterripened caryopses was inhibited by high temperature, the declining of the transcripts in afterripened embryos was stopped. The expression of these genes was also induced by ABA and water stress. Partial DNA sequence analyses indicated that some of the cDNA clones encode for LEA (late embryogenesis-abundant) proteins. To investigate whether the afterripening-induced changes in gene expression are at the transcriptional or post-transcriptional level, we chose four dormancy-associated genes to estimate their relative transcription rates and the stability of their corresponding transcripts in afterripened and dormant embryos. The transcription activities for those genes were higher to some extent in dormant embryos than in afterripened embryos 24 h after incubation, as determined by nuclear run-on assays. The half-lives of the transcripts were estimated by the use of actinomycin D. The results indicate that the half-lives of the transcripts were on the order of several hours in afterripened embryos and hundreds of hours in dormant embryos. We conclude that afterripening regulates the expression of dormancy-associated genes in excised embryos mainly at the post-transcriptional level, and transcriptional control may play a minor role.

Degree

Ph.D.

Advisors

Foley, Purdue University.

Subject Area

Botany|Molecular biology|Genetics

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