Protocol optimization for elimination of sugarcane bacilliform virus and rapid propagation of virus-free sugarcane using meristem tip culture

Tissue culture protocol was optimized for the propagation of virus-free sugarcane from infected plants using meristem tips as an explant source for the elimination of sugarcane bacilliform virus (SCBV).Virus identification on the mother (source) plant and virus indexing to monitor elimination in the tissue culture-derived plants were done by polymerase chain reaction (PCR) using degenerate SCBV primers. Murashige and Skoog (MS) media supplemented with 0.5 mg/l BAP + 0.25 mg/l kinetin and 0.1 mg/l GA(3) + 0.5 mg/l NAA were the best hormone combination for shoot multiplication and root induction, respectively. Two explant size categories (< 1 mm and 1-2 mm) were used to assess the effect of explant size on shoot regeneration and virus elimination. The results showed that explant size affects shoot regeneration. Smaller sized (< 1 mm) explants showed higher virus elimination efficiency; however, the survival frequency of explants during initiation of shoot cultures was higher in larger (1-2 mm) meristems (64.3%) in comparison to the smaller ones (35.7%). In conclusion, in vitro meristem tip culture alone is not a satisfactory approach for the generation of SCBV-free plant from infected mother plant. The virus elimination efficiency could be enhanced by using the combination of meristem tip culture with other therapies.