An investigation into the effects of Ectopic expression of transporter genes during seed development of Pisum sativum L.

Abstract

Increases in yield of food plants are urgently required. Identification of new breeding targets for genetic improvement is critical, and for which transgenic plants can provide proof-of-concept. Flower number, fruit set and seed development are crucial components of final yield in legumes and import of sucrose together with amino acids and amides largely accounts for the biomass gain and quality of the seed. Cytokinins have been implicated at all three stages of reproductive development. Besides an effect on cell division and sink establishment, the cytokinins have been implicated in the regulation of source-sink relationships. Previously developed double transgenic lines expressing both a PsAAP (an amino acid permease) and a PsSUT (a sucrose transporter) had increased seed yield due to a significant increase in seed number compared to their corresponding wild-type cultivar. This study assessed the expression of PsSUT, PsAAP, cytokinin, cell wall invertase, and SWEET gene family members during seed development in two cultivars of Pisum sativum L. Subsequently, young pods, seeds, seed coats and leaves from wild type and transgenic plants over-expressing the PsAAP and PsSUT genes were analysed to identify what effect the transgenes had had on influencing gene expression in the source and sink tissues. The genes related to cytokinin homeostasis, nutrient transport and metabolism were isolated and assessed with polymerase chain reaction (PCR) and real time-quantitative PCR (RT-qPCR) analysis. Differences were seen in the spatial and temporal expression of particular gene family members between the wild-types and their transgenic lines. In conclusion, it is evident from the data that the yield increase could be the result of enhancing both sucrose and amino acid import to the seed coat and thence to the seed, along with enhanced cytokinin and cell wall invertase activity during early stages of development. It is also evident from the data that the double transgenes had a much wider effect on the whole plant as evidenced by the enhanced gene expression in the younger source leaves.