Invitro regeneration and standardization of Agrobacterium mediated transformation of immature embryo callus from Sweet sorghum

Abstract

Abstract Sweet sorghum is the present target and cheap source for biofuel production. Tissue culture recalcitrance and low levels of transformation reproducibility of protocols are main constrains for transgenic sweet sorghum development. Production of a transient transgenic sweet sorghum for the development of disease resistant transgenics was the aim of our studies. In this study we have developed an efficient regeneration and transformation system for in vitro culture of sweet sorghum from the immature embryo callus. Histological studies of sweet sorghum embryogenic calli revealed that the development of embryogenic shoots. Murashige and Skoog nutrient agar medium with different concentrations of 2mg/l 2,4-D and combination of 0.2 mg/l NAA and 2 mg/l Kinetin gave higher frequency of callus induction and shoot regeneration. Embryogenic callus was competent to accept the DNA with GV2600 strain. These tissues are susceptible to Agrobacterium mediated transformation carrying pCAMBIA1301 with gus gene construct as well as for shoot multiplication. For effective transformation, GV2600 strain carrying pCAMBIA1301 at 0.6 O.D. was found to be compatible in giving sweet sorghum transgenics. Molecular confirmation was done by Polymerase Chain Reaction and Southern blotting for the putative plants. The regenerated plantlets survived during acclimatization were growing similar to the normal plants. All the major parameters that affecting transformation frequency were optimized for the development of perfect sweet sorghum transgenics.