Development of reproducible regeneration and transformation system for Sesamum indicum

Abstract

Sesamum indicum is an important oilseed crop with beneficiary nutrients such as essential fatty acids, proteins, and lignan. S. indicum crop improvement using conventional breeding methods was disastrous. Moreover, no reproducible regeneration and transformation protocol for large-scale generation of independent primary transformed plant lines of S. indicum virtually exists. Therefore, in the present study, a reproducible regeneration and transformation method was developed for S. indicum. Direct shoot regeneration (94.44%) was achieved using plumule tips as an explant source and when maintained on regeneration medium (BM11) an amalgamation of half the strength of MS macronutrients and B5 macronutrients together with MS micronutrients and vitamins. Maximum number of shoots (~ 16–19 shoots/explant) was obtained on BM11 medium supplemented with TDZ (1 mg/L) and BAP (0.1 mg/L). High root induction efficiency (~ 97%) was observed on RM4 medium consisted of half MS macronutrients, SH micronutrients and vitamins. The developed protocol was found effective for other commercially valuable genotypes such as Co 1, Phule til and Tilottama as well. Successful regeneration was subsequently extended to transformation of reporter gene (gfp: green fluorescent protein) as revealed by molecular analyses includes Southern hybridization, northern hybridization, real time PCR and histological study of the transformants developed in the present study. Based on the Southern analysis the calculated transformation frequency was found to be 1.33%. The presently developed regeneration and transformation system might facilitate any desired improvement of sesame crop. The present study demonstrated an improved regeneration and transformation method for S. indicum, which is genotype independent and can undergo desired modifications.