Agrobacterium tumefaciens-mediated transformation of taro (Colocasia esculenta (L.) Schott) with a rice chitinase gene for improved tolerance to a fungal pathogen Sclerotium rolfsii

Abstract

Taro (Colocasia esculenta) is one of the most important crops in the Pacific Islands, however, taro yields have been declining in Hawaii over the past 30 years partly due to diseases caused by oomycete and fungal pathogens. In this study, an efficient Agrobacterium tumefaciens-mediated transformation method for taro is first reported. In total, approximately 200 pieces (8 g) of embryogenic calluses were infected with the super-virulent A. tumefaciens strain EHA105 harboring the plant transformation plasmid pBI121/ricchi11 that contains the rice chitinase gene ricchi11. The presence and expression of the transgene ricchi11 in six independent transgenic lines was confirmed using polymerase chain reaction (PCR) and reverse transcription-PCR (RT-PCR). Southern blot analysis of the six independent lines indicated that three out of six (50%) had integrated a single copy of the transgene, and the other three lines had two or three copies of the transgene. Compared to the particle bombardment transformation of taro method, which was used in the previous studies, the Agrobacterium-mediated transformation method obtained 43-fold higher transformation efficiency. In addition, these six transgenic lines via Agrobacterium may be more effective for transgene expression as a result of single-copy or low-copy insertion of the transgene than the single line with multiple copies of the transgene via particle bombardment. In a laboratory bioassay, all six transgenic lines exhibited increased tolerance to the fungal pathogen Sclerotium rolfsii, ranging from 42 to 63% reduction in lesion expansion.