Transgenic Brassica napus L. lines carrying a two gene construct demonstrate enhanced resistance against Plutella xylostella and Sclerotinia sclerotiorum

Abstract

Plant diseases and insect pests are serious threat to the growth and yield of oilseed rape. In this study, a binary vector carrying sporamin and chitinase PjChi-1 genes in tandem was introduced into Brassica napus cv. ZS 758 via Agrobacterium tumefaciens for dual resistance against disease and insect attack. Thirty-two regenerated plantlets exhibiting hygromycin resistance were selected following Agrobacterium-mediated transformation of 600 leaf petiole explants. Of these, 27 transformants were confirmed to carry the two transgenes as detected by polymerase chain reaction (PCR) with 4.5% transformation efficiency. Eight plantlets were randomly selected for further confirmation by Southern and northern blot hybridization analyses. Four plants carried single copy of the transgenes, while the remaining four plants carried either two or three copies of the transgenes. Moreover, expression of the sporamin transgene was detected by northern blot hybridization in transgenic lines, but not in wild-type plants. These eight T 0 plants were grown in vitro, and inoculated with the Lepidoptera larvae of Plutella xylostella and with spores of the fungal pathogen of Sclerotinia sclerotiorum. Transgenic plants exhibited high levels of resistance to P. xylostella and S. sclerotiorum when compared to untransformed wild-type plants. Genetic analysis of T 1 progeny confirmed Mendelian segregation of the introduced genes. Therefore, these transgenic lines demonstrate a promising potential for variety development of oilseed rape lines with enhanced resistance against both P. xylostella and S. sclerotiorum.

Breeding for insect resistance in oilseed rape: Challenges, current knowledge and perspectives

“…Partial to complete resistance was obtained against the diamondback moth (Wang et al., ). The same result was found with a combination of chitinase and sporamin, a major protease inhibitor found in sweet potato tubers (Liu et al., ).…”

Efficient selection and evaluation of transgenic lines of Crambe abyssinica

“…If the selection pressure is too high, no transgenic lines will be recovered, while more escapes will be produced if the selection pressure is too low. A wide range of hygromycin concentrations have been used for transformation of various plant species, ranging from 3 mg L −1 for rapeseed (Liu et al, 2011; Zhang et al, 2011) to 100 mg L −1 for onion (Eady and Lister, 1998). In this study, hygromycin at 5 mg L −1 could already significantly reduce the regeneration frequency (18%) of crambe hypocotyls compared to 94% when no hygromycin was added in the medium.…”

Expression of an endochitinase gene from Trichoderma virens confers enhanced tolerance to Alternaria blight in transgenic Brassica juncea (L.) czern and coss lines

“…For example, B. napus transformed with two genes sporamin and chitinase PjChi-1 (derived from sweet potato and Paecliomyces javanicus, respectively) showed enhanced resistance against both Plutella xylostella and Sclerotinia sclerotiorum (Liu et al 2011). Wu et al (2009) introduced a plant defensin gene Ovd, cloned from Orychophragmus violaceus (L.) into B. napus, and the transgenic plants showed resistance to S. sclerotiorum.…”