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United States Department of Agriculture

Agricultural Research Service

Title: Improved Transformation Efficiency and Tissue Specific Expression of Heterologous Gs1-2/gus Genes in Maize by Utilizing the Agrobacterium Tumefaciens-Mediated Transformation Method

Authors
item Muhitch, Michael
item Sollenberger, Kurtis

Submitted to: American Society of Plant Biologists Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: July 30, 2003
Publication Date: N/A

Technical Abstract: The protein encoded by the maize glutamine synthetase GS1-2 gene is found in abundance in the maternal tissues that surround the developing maize kernel, where it is believed to play an important role in the assimilation of nitrogen released from the phloem trace. The promoter from GS1-2 is capable of directing reporter gene expression in these same maternal seed tissues after transformation of embryogenic maize calli by particle bombardment. Analysis of the parameters of transformation and expression suggest, however, that the GS1-2/GUS transgene, while readily incorporated into the maize genome, was not expressed efficiently in herbicide-resistant calli and even less so in regenerated plants. Truncation of the promoter from 664 bp to 400 bp upstream of the putative transcription start site increased expression efficiency, but resulted in the loss of tissue specificity within the kernel. In trans, the GS1-2 promoter also strongly drove GUS expression in pollen grains, despite the absence of native GS1-2 expression in pollen. Southern analysis revealed that biolistic transformation with the GS1-2/GUS gene results in high copy numbers and frequent rearrangements. We are currently transforming maize with the full length GS1-2/GUS gene construct using Agrobacterium tumefaciens-mediated transformation. Preliminary results indicate greatly increased efficiency of GS1-2/GUS transgene expression in regenerated plants, no ectopic pollen expression, and maintenance of maternal tissue specific expression within the kernel tissues. Complete analysis of transformation efficiencies, transgene copy number and expression patterns for the GS1-2/GUS gene construct in maize using Agrobacterium tumefaciens-mediated transformation will be presented.

Last Modified: 4/19/2014
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