Agrobacterium-Mediated Genetic Transformation of Brinjal with cry1F Gene for Resistance against Shoot and Fruit Borer

Abstract

Fruit and shoot borer is a major pest in brinjal (Solanum melongena L.), causing 60%–70% yield losses even after repeated insecticidal sprays. To reduce yield losses caused by this insect, transgenic plants expressing cry1F gene driven by the CaMV35S promoter were developed in four brinjal cultivars, viz., ‘Manjarigota’, ‘Ruchira’, ‘Poona selection’, and ‘Krishna kathi’. The kanamycin concentration was standardized and 50 mg/l was found to be optimum based on lethality of the explants. Cultivar ‘Ruchira’ exhibited highest (19.8%) transformation efficiency. Agrobacterium concentration of 0.2 at optical density₆₀₀ (OD₆₀₀) was found to be optimum. Highest transformation efficiency was obtained using 7-day-old shoot tip explants, which were pre-cultured for 2 days on the Murashige and Skoog (MS) medium containing 2 mg/1 benzylaminopurine (BAP) and 0.1 mg/1 indole-3-acetic acid (IAA). Explants were co-cultivated with Agrobacterium for 3 days, and then transferred to MS medium supplemented with 2 mg/1 BAP, 0.1 mg/1 IAA, 50 mg/l kanamycin, 250 mg/l cephotaxime, and 250 mg/l carbenicillin. These explants were maintained for 4–5 weeks. Rooting was done on MS medium supplemented with 1 mg/1 indole-3-butyric acid (IBA). Selected kanamycin-resistant putatively transformed brinjal lines were evaluated via polymerase chain reaction (PCR) amplification of expected 1900 bp size fragment with cry1F gene-specific primers. These transgenic lines may be effectively used for controlling fruit and shoot borer in brinjal.

KEYWORDS:

• antibiotics
• co-cultivation
• explant age
• optical density
• shoot tip
• transformation efficiency
• transgenic