Agrobacterium-mediated PsCBL and PsCIPK gene transformation to enhance salt tolerance in indica rice (Oryza sativa)

Salt tolerant crops are essential to bring the currently uncultivable salinity affected areas under cultivation. Here, we report Agrobacterium-mediated transformation of Pisum sativum calcineurin B-like protein (PsCBL) and P. sativum calcineurin B-like (CBL)-interacting protein kinase (PsCIPK) genes for development of salt-tolerant rice (Oryza sativa L.). The four indica rice cultivars used in this study were ‘BR 11’, ‘BRRI dhan 49’, ‘Paijam’, and ‘Chinigura’, with the maximum callus transformation frequency reaching 66.6 ± 7.2, 5.1 ± 6.2%, 79.1 ± 7.2 and 58.3 ± 7.2%, respectively. Among the transformed calli, the highest rates of regenerated plantlets were 62.9 ± 8.2% for ‘BR 11’, 73.5 ± 7.8% for ‘BRRI dhan 49’, and 65.5 ± 7.8% for ‘Chinigura’. ‘Paijam’ calli were recalcitrant to regeneration, which led us to directly transform ‘Paijam’ seeds without an intervening callus phase. The maximum transformation rate of ‘Paijam’ plantlets obtained was 46.3 ± 5.3%. Successful transformation events were confirmed through GUS histochemical assays and PCR analyses of putative transgenic plants. The transgenic rice plants ‘BR 11’, ‘BRRI dhan 49’, ‘Paijam’, and ‘Chinigura’ were able to survive exposure to 150 mM NaCl stress for over 3 wk, while non-transformed control plants became bleached and died within 1 wk for the same treatment. Our results indicated that stable transformation of these rice cultivars either with PsCBL or PsCIPK significantly improved salt tolerance.