Construction of ihpRNA Expression Vector of MsLEA3-1 from Medicago sativa L. and Genetic Transformation in Tobacco

Abstract

Late embriogenesis abundant (LEA) protein is one of the hot topics in plant stress physiology. In this study, an RNAi expression vector harboring MsLEA3-1 gene fragment from alfalfa (Medicago sativa L.) was constructed. Two pairs of specific primers with different enzyme sites were designed based on the sequence of MsLEA3-1 (GenBank accession number EU665182). With the template of PMD-LEA plasmid constructed, positive-sense strand and antisense strand were obtained, which were separately inserted into the expression vector pART27. A hairpin structure in the RNAi vector pART-F-R was confirmed by the digestion of restriction enzymes. pART-F-R was transformed into tobacco via Agrobacterium-mediated transformation system, and 16 transgenic plants were obtained after PCR validation.