Evaluation of a new transformation method in tomato

Abstract

A new system for transformation of tomato was evaluated using the Helios Gene Gun. Transformation parameters were optimized using a plasmid containing the GUS gene. The bombarded leaflets were subjected to the GUS histochemical assay and the number of blue foci was counted. The highest GUS expression was obtained when using helium pressure of 200 psi with 0.6um gold particle size. The microcarrier loading quantity (MLQ) of 0.25 mg/shot was found to be the most convenient for use with the Helios Gene Gun. A promoter comparison study was carried out with both the Helios Gene Gun and the Biolistic Gun. The level of expression of the promoters in driving the GUS expression was estimated by counting the number of blue foci expressed in the bombarded leaflets. The CsVMV promoter showed the highest expression followed by the CaMV 35$ promoter then the Maize Ubil promoter while the Rice Actl showed the lowest average expression. Similar results were obtained with both the Biolistic Gun and the Helios Gene Gun. Apical meristems of 12 plants from each of the 3 cultivars (Castlerock, Strain B and Peto 86) were bombarded with the construct harboring the CsVMV promoter using the Helios Gene Gun. Transformed plants were screened for putative transgenic events using PCR. No amplification products were obtained revealing no sign of stable transformation. The effects of the new system on the morphological characters of the bombarded plants were investigated. Statistical analysis of data collected on different morphological characters indicated no significant differences between the bombarded and non• bombarded plants using the Helios Gene Gun.