Development and Molecular Characterization of Transgenic Pigeon Pea (Cajanus cajan [L.] Millsp.) Plants Overexpressing OsLecRLK Using Agrobacterium Meditated Genetic Transformation for Salinity Stress Tolerance

Abstract

Aim: Pigeon pea productivity is affected by various types of abiotic stresses such as salinity, drought, high and low temperatures etc. Salt stress has become the major environmental constrain which leads to yield loses. Therefore, the development of salt tolerant varieties is the sole need of time and engineering crops to attain their maximum potential is the best option. Methodology: In the present study, transgenic pigeon pea (cv. Manak) plants carrying OsLecRLK gene, has been developed for salinity tolerance by using a robust and efficient transformation protocol, called inplanta transformation protocol. Agrobacterium strain LBA4404 harbouring pCAMBIA1301 containing OsLecRLK gene was used for the transformation of pigeon pea (cv. Manak) using Agrobacterium-mediated transformation protocol for which patent has already been granted. Results: Putative transformants were screened through PCR amplification using gene specific primers and 16 plants out of 86 plants were found positive giving a transformation efficiency of 18.6%. Higher yielding lines were selected and checked for stable transgene integration. Southern hybridization and Real-time PCR analysis was done to find out the copy no. of transgene in selected transgenic lines. Seeds from T0 pigeon pea plants were sown to raise T1 generation and further screened for the presence of OsLecRLK gene. Conclusion: Transgenic pigeon pea plants were produced harbouring OsLecRLK gene with a rapid and efficient non tissue culture based transformation method with an efficiency of 18.6 %. This method can be used for transformation in other modal plants. Five out of six selected lines confirm single copy insertion through Southern Blotting and Real Time PCR, which is highly recommended in case of transgenic plants.