I. A. Hafiz

Assessing the response of indigenous loquat cultivar Mardan to phytohormones for in vitro shoot proliferation and rooting

In vitro cultures of loquat cultivar Mardan were established using shoot apices after treating with NaOCl (5%, 7%, 10%, 12%, 14% (v/v)) for 12 min and HgCl2 (0.01%, 0.05%, 0.10%, 0.20%, 0.25% (w/v)) for 2 min. A maximum survival rate of 70% was recorded after surface sterilization with 10% NaOCl. Caulogenic response was assessed on Murashige and Skoog (MS) medium fortified with assorted combinations of the cytokinins, benzylaminopurine (BAP), kinetin, and N6-(2-isopentyl)adenine (2iP). Treatment of BAP 1.5 mg/L combined with 2iP 9.0 mg/L and kinetin 1.5 mg/L was found to be optimum for shoot morphogenesis in terms of the number and subsequent growth of shoots, while the highest shoot length was yielded by the combination of BAP 0.5 mg/L, kinetin 0.5 mg/L, and 2iP 3 mg/L. Higher levels of cytokinins induced callogenesis, vitrification and stunted growth to some extent. For rhizogenesis, uniform sized micro-shoots were excised and transferred to half-strength MS medium containing auxins. The best rooting expression was observed with naphthaleneacetic acid (NAA) 1 mg/L combined with indole-3-butyric acid (IBA) 2 mg/L and paclobutrazol (PBZ) 1 mg/L.

Effect of Various Factors on the Efficiency of Agrobacterium-Mediated Transformation of Grape (Vitis vinifera L.)

The present investigation was conducted to study the effect of inoculation time, cocultivation period and various concentrations of hygromycin and kanamycin on genetic transformation of grape (Vitis vinifera L.) cv. Kings Ruby through Agrobacterium tumefaciens. The Chitinase gene (for fungal resistance) and GUS gene (for phenotypic expression of transgenic) were introduced in the embryogenic calli developed from leaf discs, through Agrobacterium strain LBA4404 harboring plasmid pBI121 nptII as selectable marker for GUS gene and hptII for Chitinase gene. Regarding transformation efficiency rate, 10 minutes inoculation period with bacterial suspension showed the highest transformation efficiency rate i.e. 2.83% with Chitinase gene and 2.5% with GUS gene. Infected calli with Chitinase gene and GUS gene co-cultivated for 2 days showed the maximum transformation efficiency of 2.75% and 3.25%, respectively. For elimination of excess bacteria, cefotaxime treatment (300 mg L−l) showed the highest survival rate of 3.16% for Chitinase gene and 2.5% for GUS gene. The maximum i.e.2.83% transformation efficiency rate was achieved at 10 mg L−l of hygromycin for selection of transformed calli. The highest 2.25% transformation efficiency was yielded when kanamycin was used at the rate of 100 mg L−l. Five out of 8 calli showed positive expression with 62.5% transformation efficiency through histochemical GUS assay. Present results may be helpful in improving genetic transformation efficiency of grape through Agrobacterium tumefaciens against fungal diseases and reduce the use of fungicides.