Genetic engineering of maize towards desiccation tolerance: Electroporation with the trehalose gene

Abstract

Trehalose is a non-reducing disaccharide of glucose that occurs in a large number of organisms such as bacteria, fungi, nematodes or crustaceans. Trehalose plays an important role in desiccation and heat stress protection since it has been shown to stabilize proteins and cell membranes under these stress conditions. Trehalose accumulation has proven to be an effective way of increasing drought tolerance in both model plants such as tobacco and important crops such as potato or rice. In this work we aim to improve desiccation tolerance in maize, one of the, most agronomical important crops, by increasing trehalose accumulation through transformation with the Arabidopsis thaliana trehalose phosphate synthase gene (AtTPSl) is involved in trehaIose-6-phosphate synthase and hence on trehalose biosynthesis. A cassette harboring the AtTPSl gene under the control of the CaMV35S promoter and the Bialaphos resistance gene Bar as a selective agent (conferring resistance to the PPT) was inserted in the plasmid vector pGreenO229 and used to transform maize inbred line Pa91. Immature zygotic embryos were collected 14-20 days after pollination and embryogenic calli culture were initiated. Embryog?nie calli were electroporated with 20?g of plasmid DNA using a Biorad Gene Puiser II at 374 V, for 1 second. Embryogenic calli were electroporated and selected PPT. Eighty putative transgenic plants were obtained and analyzed by PCR for the presence of the AtTPS1 gene.