OPTIMIZATION OF AN EFFICIENT REGENERATION AND TRANSFORMATION PROTOCOL FOR BLACK RICE CHAKHAO POIREITON
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Date
2019-07
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AAU, Jorhat
Abstract
The black rice germplasm suffers from inherent low productivity due to inefficient performance of yield attributing characters. Besides, the crop is also susceptible to biotic and abiotic stress. Gene technology can provide an effective approach for improvement of black rice. The availability of a robust and reproducible plant regeneration system that is amenable to transformation plays a major role in achieving this objective. Therefore a study was conducted to establish a stable and reproducible regeneration system in black rice Chakhao poireiton. The main plant hormone responsible for callus induction 2,4-D was used in combination with vitamin assay casamino acid in 2N6 medium with varying concentrations to establish a better callus inducing medium. It was revealed that a combination of 2mg/l 2,4-D and 1g/l vitamin assay casamino acid stood better in terms of callus induction frequency, callus weight and relative growth rate compared to the already established protocol by Hiei et al., (2008). The best shoot inducing hormone combination was found out by using kinetin in varying concentration and kinetin in combination with BAP. Kinetin at 4.5mg/l proved to have the highest regeneration percentage. Agrobacterium strain LBA4404 harbouring pCAMBIA1304 binary vector containing hptII gene (selectable marker gene) and gusA gene (reporter gene) was used for genetic transformation. GUS histochemical Assay showed visual confirmation of genetic transformation. Varying concentrations of hygromycin 30mg, 40mg, 50mg and 60mg/l were tried for screening transformants, but gave 100% mortality in all the concentrations. In future, better transformation protocols for Chakhao poireiton need to be established.
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