Development of transgenic barstar lines and identification of a male sterile (barnase)/restorer (barstar) combination for heterosis breeding in Indian oilseed mustard (Brassica juncea)

Abstract

Transgenic lines containing the barstar gene (encoding for Barstar an intracellular inhibitor of the ribonuclease, Barnase both from Bacillus amyloliquefaciens) have been developed in Indian oilseed mustard, Brassica juncea, to develop a complete male sterility/ restoration system for heterosis breeding in this crop. Transgenics were also raised using a modified sequence of the barstar gene based on parameters known to influence transgene expression in heterologous systems. The wild type and modified barstar lines were analysed for their restoration capabilities by crossing them with agronomically suitable male sterile barnase lines developed earlier in our laboratory. Of 30 different combinations of crosses tested between three male sterile barnase lines and several single-copy barstar lines, only one combination was found to restore male fertility among F1 progeny. Subsequent analysis of F2 progeny derived from such F1 restored events (containing both barnase and barstar genes) revealed stable inheritance of both genes in the segregating population thereby indicating proper functionality of the same. Further, pollen viability in restored events was found to be comparable to that observed in transgenic lines containing the barstar gene alone, indicating efficient restoration by the barstar protein in the presence of the ribonuclease. The male sterile line and its corresponding restorer identified in the present study constitute a complete, functional male sterility/restorer system in B. juncea and the traits can be diversified into appropriate combiners for heterosis breeding.