Gene delivery into the plastome was initially done by Agrobacterium mediated method [Block et al.1985 ] [69] . The discovery of biolistic DNA delivery led to plastid trans-formation via particle gun [Sanford 1990 ] [70] . In this method, the Escherichia coli plasmids contain a marker gene and the gene of interest is introduced into plastids. The foreign genes are inserted into plasmid DNA by homologous recombination via the flanking sequences at the insertion site [66] . Polyethylene glycol (PEG) mediated transformation of plastids was also utilized [71, 72] . PEG-mediated transformation of plastids requires enzymatically removing the cell wall to obtain protoplasts, then exposing the protoplasts to purified DNA in the presence of PEG. The protoplasts first shrink in the presence of PEG, then lyse due to disintegration of the cell membrane. Removing PEG before the membrane is irreversibly damaged reverses the process. Biolistic delivery is the routine system for most laboratories, as manipulation of leaves, cotyledons, or cultured cells in tissue culture is a simple practice than the alternative PEG treatment of protoplasts [58] . Recently, particle gun mediated plastid transformation has been demonstrated in rapeseed using cotyledons as explants [38] .