Low levels of expression of wild type Bacillus thuringiensis var. kurstaki cryIA (c) sequences in transgenic walnut somatic embryos

Abstract

Insecticidal crystal protein fragments (ICPFs) of Bacillus thuringiensis encoded by cryIA (c) and cryIA (b) were previously shown to be lethal to key target walnut insect pests (codling moth, navel orangeworm and Indianmeal moth). One of these genes, cryIA (c), was used to transform walnut somatic embryos using a binary vector (pWB139) in which this gene was expressed as a protein fusion with the kanamycin resistance gene from bacteria. Transgenic embryos representing individual transformation events were germinated to produce shoots that were maintained as micropropagated shoot lines in the laboratory. Transgenic shoots were then grafted onto seedling rootstocks and transplanted into the field. DNA analysis (Southern blotting) demonstrated that these shoots were transformed by pWB139 or by the control construct pWB149 (which does not contain the cryIA (c) gene). Insect feeding trials of walnut embryos grown in vitro and infested with neonatal Indianmeal moth, codling moth or navel orangeworm larvae showed non-significant mortality; confirming insufficient or incomplete expression of ICPs in the transgenic embryos. Therefore, transformation of somatic walnut embryos with the vector pWB139 was ineffective in protecting the embryos from damage by lepidopteran insect larvae.