Development of Genetic Transformation of Peanut Mediated by Agrobacterium

Abstract

One of the constrain in peanut production is the disease due to peanut stripe virus. Since there is no source of gene for resistance to PStV among cultivated peanut, peanut cultivar with resistance to PStV is not available. Genetic transformation to introduce PStV CP gene can be used to generate cultivar with resistance to PStV. The objective of this experiment was to develop genetic transformation protocol, for various commercial cultivars of peanut from Indonesia, mediated by Agrobacterium. The first step toward developing genetic transformation protocols, mediated by Agrobacterium for peanut was the development of embryonic callus lines suitable for target materials. The embryonic callus lines were developed from mature embryo of peanut cv. Gajah. The callus lines have been maintained for almost six months and retained their capabilities for regenerating somatic embryos in vitro. The second step was the development of conditions suitable for transferring genes into the embryonic callus lines of peanut. Peanut callus lines were dipped on the culture of Agrobacterium and co-cultivated for 24 hours. After co-cultivation, the callus lines were transferred onto medium for inducing somatic embryo, containing cefotaxime (250 mg/l) to kill the Agrobacterium and kanamycin (100 mg/l) to select the transformed cells. The regenerated somatic embryos that were kanamycin resistant were germinated on germination medium with kanamycin (100 mg/l) to regenerate plantlets. The seedlings were then transferred onto soil medium and transferred into green house. Molecular and biochemical characterisations were conducted for the putative transgenic peanut.