Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 9/30/2005
Publication Date: 2/13/2006
Citation: Jia, Y., Anderson, J.V., Horvath, D.P., Gu, Y.Q., Lym, R.G., Chao, W.S. 2006. Subtractive cDNA libraries identify differentially-expressed genes in dormant and growing buds of leafy spurge (Euphorbia esula). [Abstract]. Weed Science Society of America Abstracts 46:30. Interpretive Summary: A genomics approach was applied to identify and clone genes associated with dormancy and growth in the root and crown buds of leafy spurge. Two subtracted (a forward and a reverse) cDNA libraries were developed for this work. The forward library contains genes preferentially expressed in growing buds and the reverse library contains genes preferentially expressed in dormant buds. After library screening, 521 non-overlapping genes were obtained. The expression of these genes was examined based on macroarray and semi-quantitative RT-PCR analysis. Many differentially-regulated genes during bud development were identified. Some of these genes have unknown or hypothetical functions while others are known to play important roles in molecular functions such as cytochrome P450 and lipid transfer protein.
Technical Abstract: Two subtracted cDNA libraries (growing and paradormant) were developed to study genes associated with bud dormancy and initiation of shoot growth in leafy spurge. Initial analysis revealed that both libraries contained many redundant clones. To identify unique sequences represented in each library, 15744 clones from both libraries were spotted onto membranes and sequentially hybridized with sets of clones known to be redundant within the libraries. A total of 521 unique sequences were obtained from 2304 minimally redundant clones. DNA inserts from the 521 individual clones were PCR-amplified and gel-purified prior to being used to conduct macroarray analysis. Radioactive probes developed from RNAs extracted from crown buds of either intact (control) or a series of induced (2 hr, 2 d, and 4 d after decapitation) plants were used to identify differentially-expressed genes. Semi-quantitative RT-PCR was used to confirm macroarray results and to determine the expression profiles for other transcripts from the subtracted libraries. Gene expression of selected unique sequences was also examined in crown buds after growth induction and/or during normal seasonal growth. Many of the differentially-regulated genes identified in this study have unknown or hypothetical functions while others are known to play important roles in molecular functions such as cytochrome P450 and lipid transfer protein.