|Samac, Deborah - Debby|
Submitted to: Central Alfalfa Improvement Conference
Publication Type: Abstract Only
Publication Acceptance Date: 7/26/1999
Publication Date: N/A
Technical Abstract: Alfalfa (Medicago sativa L.) suffers greatly from Al stress in acid soils, and genetic diversity for Al tolerance is very limited. Exudation of organic acids by Al-tolerant crops in response to Al exposure is very well documented. Manipulating the expression of enzymes involved in the biosynthesis of organic acids may be a good approach to enhancing Al tolerance in alfalfa. The full-length cDNAs encoding the nodule-enhanced forms of phosphoenolpyruvate carboxylase (PEPC) and malate dehydrogenase (MDH) were cloned separately in plant transformation vectors to generate 35S/neMDH, 35S/nePEPC, and ScBv/neMDH constructs. Several kanamycin- resistant primary transformants were obtained which did not show any phenotypic differences from the nontransformed Regen SY alfalfa. PCR was used to further identify transgenic lines among the kanamycin-resistant lines. In vitro enzyme activity assays and immunodetection of neMDH and nePEPC proteins on western blots were carried out and identified several lines with enhanced protein and enzymatic activity over control nontransformed plants. Growing root tips of transgenic plants acidified the rhizosphere as detected by an indicator dye, bromocresol purple. Selected transgenic lines were also shown to have increased malate concentration in root tips. Work is currently underway to characterize selected lines for PEPC and MDH mRNA accumulation. Experiments to evaluate Al tolerance and to measure nodulation and biological N-fixation should determine the potential utility of this molecular approach towards enhancing Al tolerance in crop plants.