|Samac, Deborah - Debby|
Submitted to: Plant Physiology Supplement
Publication Type: Abstract Only
Publication Acceptance Date: 6/1/1996
Publication Date: N/A
Technical Abstract: Asparagine synthetase (AS) catalyzes the glutamine-dependent amidation of aspartate, producing asparagine. An 8 kb genomic clone containing an entire AS gene was isolated and sequenced. Sequencing of more than 20 cDNA clones from both root and nodule libraries showed a single cDNA species with varying polyadenylation sites. Gel blot hybridization to RNA isolated dfrom various alfalfa tissues shows that the level of AS mRNA is high in nodules but low in leaves, stems, roots, and cotyledons. Western blots probed with AS antisera to total protein extracted from various tissues reveal that the amount of AS protein is approximately 50-fold higher in nodules than in leaves, stems, meristems, roots, or cotyledons. Increases in both AS mRNA abundance and AS protein amount in leaves and roots were detected following dark treatment. A 2.7 kb 5'-flanking region of the gene fused to the Beta-glucuronidase (GUS) reporter gene was transformed into alfalfa to examine the regulation of the AS gene. Analysis of transgenic alfalfa shows that GUS activity is mainly detected in symbiotic zones of the nodules. In situ hybridization showed that AS was highly expressed throughout the symbiotic zone of effective nodules while expression in ineffective nodules was limited to only a few layers of infected cells.