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Cloning of PA1 gene from Pisum sativum and genetic transformation on Medicago sativa

ZHANG Gai-na1,2,JIA Jing-fen2(1.College of Agriculture of Henan University of Science and Technology,Luoyang 471000,China;2.School of Life Science,Northwest University,Xi'an 710069,China)  
A gene,a sulphur-rich and insect-toxic seed albumin-PA1 gene,was cloned by PCR method from Pisum sativum(cultivar shijiadacaiwan).In order to characterize the function of this gene,a plant expression vector,pCAMBIA1301-PA1,was constructed.This vector was transformed into M.sativa via Agrobacterium-mediated method.The conditions for transformation were optimized.The several transgenic embryogenic calli and regenerated plantlets were obtained.PCR and Southern blotting identification confirmed that PA1 gene and hygromycin-B phosphotransferase-resistant gene has been integrated into the genome of M.sativa,SDS-PAGE analysis indicated that PA1 gene expressed in the transgenic lines.Free amino acid analysis showed that the levels of methionine and cysteine in tansgenic M.sativa.Plantlets improved from 0.1% to 0.4%.
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