Abstract
Agrobacterium mediated transformation has been widely used for research in plant molecular biology and for genetic improvement of crops exploiting its tremendous ability to transfer foreign DNA to plants. In this study, the transformation efficiency of five Agrobacterium tumefaciens strains GV2260, LBA4404, AGL1, EHA105, and C58C1 was evaluated in Nicotiana tabacum L. cultivar Samsun. The Agrobacterium strains contained therecombinant binary vector pBin19 harboring beta-glucuronidase uidA gene under 35S promoter. Neomycinphosphotransferase (nptII) gene was used as a selectable marker at a concentration of 100 mg L-1 kanamycin. The expression of uidA gene in regenerated T0 plants was firstly analyzed by GUS histochemical analyses andlater on confirmation of presence of the nptII and uidA genes in regenerated plants was determined by PCR. The highest transformation rate (20%) was obtained with the Agrobacterium strain LBA4404, followed by EHA105, GV2260, C58C1 and AGL1. The higher transformation efficiency achieved in our studies make LBA4404 Agrobacterium strain optimal for functional genomics and biotechnological applications in tobacco plants.
Highlights
Plant transformation employs the use of Agrobacterium tumefaciens in most of the cases which is a naturally occurring soil borne pathogenic bacterium that causes crown gall disease
Agrobacterium tumefaciens infects the wound sites in dicotyledonous plant resulting in the formation of the crown gall tumors
The crown gall disease is due to the transfer of a specific fragment “T-DNA” from a large tumor-inducing (Ti) plasmid within the bacterium to the plant cell (Zaenen et al, 1974)
Summary
Plant transformation employs the use of Agrobacterium tumefaciens in most of the cases which is a naturally occurring soil borne pathogenic bacterium that causes crown gall disease. The present studies was conducted to evaluate the transformation efficiency of five Agrobacterium tumefaciens strains, GV2260, LBA4404, AGL1, EHA105, C58C1, harboring the plasmid pBin19 containing beta-glucuronidase uidA gene under 35S CaMV promoter (Figure 1). Culture was spread on plates containing LB medium and 50 mg/L of kanamycin for selection of transformed cells.
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