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United States Department of Agriculture

Agricultural Research Service

Title: Alfalfa Aspartate Aminotransferase-2: in Situ Expression of Mrna in Root Nodules and Analysis of the Putative Promoter Region of the Gene

item Yoshioka, Hirofumi - UNIVERSITY OF MINNESOTA
item Gregerson, Robert - UNIVERSITY OF MINNESOTA
item Samac, Deborah
item Vance, Carroll

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: July 14, 1996
Publication Date: N/A

Technical Abstract: Aspartate aminotransferase isoform-2 (AAT-2) plays a key role in the assimilation of ammonia from symbiotic nitrogen fixation into amino acids. We previously documented the characterization of the AAT-2 cDNA, AAT-2 mRNA expression in various tissues with RNA blots, AAT-2 protein synthesis, and the structure of the AAT-2 gene. To further understand the genetic regulation of this key enzyme, we have assessed the pattern of AAT-2 gene expression within cells of effective and ineffective root nodules of alfalfa through in situ hybridization. We have also evaluated the 5' upstream putative promoter region of AAT-2 in transgenic alfalfa transformed with promoter deletions fused to B-glucuronidase (GUS). In situ hybridizations showed that AAT-2 expression in elongate alfalfa nodules that have indeterminate meristems occurs throughout Zone III of effective nodules. Expression appears to be initiated prior to detectable nitrogenase activity and is fully detectable in the first cell layer dista to the prefixing zone. AAT-2 message is much higher in bacteroid containing cells than any other cell type. In transgenic alfalfa, a chimeric reporter gene containing 2500 bp of the 5' upstream region of AAT- 2 fused to GUS was expressed in both infected and uninfected cells of Zone III and slightly in nodule parenchyma. Analyses of promoter deletions showed that the region between -410 and -147 was important in directing transcriptional activity to the infected cell zone (Zone III) of alfalfa nodules. Similar results were obtained when the promoter constructs were transformed into Lotus corniculatus. Thus, the promoter region of AAT-2 from alfalfa shows fidelity of expression in a heterologous host species having spherical nodules and a determinate meristem.

Last Modified: 4/18/2015
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