A reliable and high-efficiency Agrobacterium tumefaciens-mediated transformation system of Pogonatherum paniceum embryogenic callus using GFP as a reporter gene
Successful efforts describing in vitro culturing, regeneration, and transformation of grain sorghum were reported, using particle bombardment, as early as 1993, and with Agrobacterium tumefaciens in 2000. Reported transformation efficiencies via Agrobacterium routinely range from 1 to 2%. Recently, such efficiencies via Agrobacterium in several plant species were improved with the use of heat and centrifugation treatments of explants prior to infection. Here, we describe the successful use of heat pretreatment of immature embryos (IEs) prior to Agrobacterium inoculation to increase routine transformation frequencies of a single genotype, P898012, to greater than 7%. This reproducible frequency was calculated as numbers of independently transformed IEs, confirmed by PCR, western, and DNA hybridization analysis, that produced fertile transgenic plants, divided by total numbers of infected IEs.