Agrobacterium-mediated transformation of Brassica rapa with a bacterial isochorismate synthase (ICS) gene

The entC gene from Escherichia coli which codes for isochorismate synthase (ICS) was introduced into Brassica rapa varieties i.e. var. rapa cv. Raapstelen and ssp. oleifera. Transgenic plants of ssp. oleifera carrying the ICS gene have been developed via Agrobacterium tumefaciens-mediated transformation. This method involved the use of hypocotyl explants co-cultivated with disarmed Agrobacterium strain LBA4404, which on the T-DNA region carries CaMV 35S promoter driven genes ssentC, consisting of the entC coding region fused to the coding region of the plastid targeting sequence of tobacco ribulose-bisphospate carboxylase, hptII, encoding hygromycin phosphotransferase, and a gene for sGFP. Prior to this, several important parameters were examined for their influence on the transformation efficiency. The presence of the transgenes in the genome of the nine months old greenhouse-grown transgenic plants was confirmed by PCR, whereas the expression of the entC gene was revealed by RT-PCR. The results indicated that the exogenous gene was successfully integrated into the genome and expressed in transgenic B. rapa plants.