Isolation and characterization of a harvest-inducible gene hi11 and its promoter from alfalfa
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The harvesting and storing of alfalfa is a routine practice in the agricultural industry worldwide. To investigate gene expression in harvested alfalfa, cDNA from non-harvested and harvested plants in the field was subjected to subtractive hybridization to identify, in particular, those genes that are induced by the harvesting treatment. One cDNA clone, named hi11, was isolated and analysed. The full length cDNA of the hi11 gene was cloned by RACE amplification. The hi11 gene, which has high homology to a putative protein of unknown function in Arabidopsis, was induced in alfalfa following harvesting, a 38°C heat shock and a wounding treatment. Northern blot analysis confirmed that the expression patterns of hi11 in alfalfa in response to harvesting, heat shock, and wounding. In addition, genomic walking was performed to isolate the 5′ flanking region of the hi11 gene. The promoter of the hi11 gene was fused to the GUS reporter gene and transferred to Medicago truncatula and tobacco. In all transgenic plants of M. truncatula and tobacco, GUS gene expression was observed in harvested tissue, especially in the transgenic tobacco plants, but not in the non-harvested control tissue.
KeywordsHarvest-induced gene Subtractive hybridization RACE Genome walking hi11 Alfalfa
The authors gratefully acknowledge support for this research from the Canadian Natural Sciences and Engineering Research Council, Strategic Grants Program.
- 2.Barnes DK, Goplen BP, Baylor JE (1988) “Highlights in the USA and Canada” in alfalfa and alfalfa improvement. American Society of Agronomy, Inc., Madison, pp 1–22Google Scholar
- 12.Trieu AT, Burleigh SH, Kardailsky IV, Maldonado-Mendoza IE, Versaw WK, Blaylock LA, Shin H, Chiou TJ, Katagi H, Dewbre GR, Weigel D, Harrison MJ (2000) Transformation of Medicago truncatula via infiltration of seedlings or flowering plants with Agrobacterium. Plant J 22:531–541CrossRefPubMedGoogle Scholar
- 13.Sambrook J, Russell DW (2001) Molecular cloning; a laboratory manual. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
- 16.Shen X, Martens S, Chen M, Li D, Dong J, Wang T (2010) Cloning and characterization of a functional flavanone-3ss-hydroxylase gene from Medicago truncatula. Mol Biol Rep. doi: 10.1007/s11033-009-9913-8