PJB-2009-282
STUDY OF THE FACTORS AFFECTING AGROBACTERIUM MEDIATED GENE TRANSFORMATION IN TOMATO (LYCOPERSICON ESCULENTUM MILL.) CV. RIOGRANDE USING RICE CHITINASE (CHT-3) GENE
NYLA JABEEN1*, BUSHRA MIRZA1, ZUBEDA CHAUDHARY2, HAMID RASHID3 AND MUHAMMAD GULFRAZ4
Abstract
The factors affecting the Agrobacterium mediated gene transformation of tomato cv. Riogrande are reported in detail. Shoot tips (explants) were transformed with pBI333 carrying rice chitinase (cht-3) gene. Approximately 49% transformation frequency was achieved. The optimum conditions for successful transformation were: infecting the explants with the Agrobacterium suspension for two minutes, co-cultivation period for two days, 50 µM of acetosyringone in the co- cultivation medium, pre- selection for seven days and 500 mg/l cefaxine in the pre- selection. The transgenic plants were selected on the medium containing 50 mg/l hygromycin. The presence of hygromycin and chitinase gene in the hygromycin resistant (T0) transgenic plants was evaluated by Polymerase Chain Reaction (PCR) analysis. A fragment of 500bp size for the hygromycin gene was amplified from the 100% hyg resistant plants while 250 bp long fragment for the cht-3 gene was amplified from the 88% of the transgenic plants only.
To Cite this article:
Download
