Agrobacterium-mediated Genetic Transformation of two Peanut (Arachis hypogaea L.) Varieties using Salinity Tolerant PDH45 Gene

Abstract

An efficient Agrobacterium-mediated genetic transformation protocol has been developed in the present investigation to integrate salinity tolerant gene pea DNA helicase (PDH45) in two peanut varieties, Dhaka-1 and BARI Chinabadam-8. Decapitated half-embryo explants of the above-mentioned varieties were also used to develop a transformation compatible in vitro regeneration system. MS medium supplemented with 5.0 mg/l BAP and 0.5 mg/l Kn showed maximum response for shoot induction in both varieties. Half strength of MS medium supplemented with 0.2 mg/l IBA showed better response towards root induction for both varieties. Genetic transformation experiments were performed using Agrobacterium strain LBA4404 containing pCAMBIA1301-PDH45 conferring reporter gus gene, hygromycin resistance hpt gene, and salinity tolerant gene pdh45. The transformation variables were optimized, OD₆₀₀ 1.4 with 45 minutes of incubation period, and three days of co-culture showed the best performance. The transformed shoots were selected using 30 mg/l hygromycin. The overall transformation frequency was very low, where 0.10% shoots of Dhaka-1 and 0.07% shoots of BARI Chinabadam-8 survived the selection pressure. The stable integration of the transgene in the transformed shoots was confirmed through PCR analysis.

Plant Tissue Cult. & Biotech. 33(1): 37-46, 2023 (June)