Optimization of an integrated bombardment and agrobacterium infiltration based transformation system for lentil

Çelikkol, Ufuk
In this study, the effect of microwounding by particle bombardment and the coupling of vacuum infiltration to Agrobacteritim-mediated transformation, on the efficiency of transient GUS expression of cotyledonary node tissues were investigated. In each experimental set with any Agrobacterium/pia&mid couple, five different treatments were applied to compare the efficiency of integrated bombardment-infiltration system to reference treatments. Throughout the study also the efficiencies of four strains, LBA4404, EHA105, C58C1, KYRT1 and two binary plasmids, pGUSINT and HIpTJK136 on transient GUS expression of cotyledonary node explants were compared. Although differ upon inoculation of different strain/binary plasmid couples, overall transient expression appeared to be reduced when tissues were microwounded. Furthermore, no significant (p>Q.0S) effect of the vacuum infiltration was observed. Therefore, efficiency of another wounding method, mechanical injury (poking by a needle) of the cotyledonary petiole axil region was investigated. Usage of the most efficient strain/plasmid couple, which is determined to be KYRTl:pTJK136, and mechanical wounding of the cotyledonary petiole axil region of the explants prior to Agrobacterium inoculation produced a number of transformed shoots, which show stable GUS expression in a chimeric manner. Highly regenerable nature of the cotyledonary node explants and application of the above mentioned transformation protocol seem to increase the probability of obtaining transgenic plants in lentil.


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In this study, optimization of a transformation and regeneration system via indirect organogenesis in cotyledonary petiole tissue of lentil (Lens culinaris Medik.) was investigated. Eight different medium types differing in their plant growth regulator compositions were employed to examine the callus induction potency of cotyledonary petiole. Except two, all other tested medium yielded more than 80% callus induction. Nine different medium types were studied to test the potencies of callus structures for sho...
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Citation Formats
U. Çelikkol, “Optimization of an integrated bombardment and agrobacterium infiltration based transformation system for lentil,” Middle East Technical University, 2002.