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STUDY OF THE FACTORS AFFECTING AGROBACTERIUM MEDIATED GENE TRANSFORMATION IN TOMATO (LYCOPERSICON ESCULENTUM MILL.) CV. RIOGRANDE USING RICE CHITINASE (CHT-3) GENE

NYLA JABEEN^1*, 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.

¹Department of Biochemistry, Quaid-i-Azam University, Islamabad,

²Department of Botany, Hazara University, Mansehra, Pakistan,

³Department of Bioinformatics, Mohammad Ali Jinnah University, Islamabad, Pakistan,

⁴Pir Mehar Ali Shah, Arid Agriculture University, Rawalpindi, Pakistan

^*Corresponding author email: nylajabeensatti@yahoo.co.in