APPLICATION OF LASER MICROBEAM CELL SURGERY AND AGROBACTERIUM–MEDIATED GENE TRANSFORMATION SYSTEMS IN MELON (CUCUMIS MELO L)

Abstract

In the current investigation, the application of laser microbeam cell surgery as a transformation system in melon has been studied and compared with transformation via Agrobacterium tumefaciens as well as a combine of the two systems. All these transformation treatments have been carried out on the hypocotyl proximal zone of melon cv. Shahd El-Dokki using the plasmid pISV2456 that harboring gus-intron and bar genes. The treated explants were regenerated on shoot regeneration medium supplemented with 250 μg/l bialaphos. Results showed that there was no significant difference of shoot regeneration between the laser and Agrobacterium treatments, however, the regeneration percentage varied among the treatments, the laser microbeam treatment showed highest percentage than the Agrobacterium and combined methods, (76, 72 and 68 % respectively). The transformation systems were evaluated by detecting the expression of the gus gene using histochemical assay, while the integration of the both genes (gus and bar) were confirmed by the PCR assays, indicating that the two genes were successfully transferred to the plants in the three methods. Seven shoots out of 30 tested shoots were PCR-positive, representing 23.33%, in both Agrobacterium and laser treatments, while, only 6 out of 30 tested shoots were PCR-positive, representing, 20%. This study demonstrates using laser tool as a new transformation method in melon cv Shahd El-DOkk, indicating that laser technique could be considered a competitive transformation method with the Agrobacterium-system.