Development of transgenic sorghum for insect resistance against the spotted stem borer (Chilo partellus)

Abstract

Transgenic sorghum plants expressing a synthetic cry1Ac gene from Bacillus thuringiensis (Bt) under the control of a wound-inducible promoter from the maize protease inhibitor gene (mpiC1) were produced via particle bombardment of shoot apices. Plants were regenerated from the transformed shoot apices via direct somatic embryogenesis with an intermittent three-step selection strategy using the herbicide Basta. Molecular characterisation based on polymerase chain reaction and Southern blot analysis revealed multiple insertions of the cry1Ac gene in five plants from three independent transformation events. Inheritance and expression of the Bt gene was confirmed in T(1) plants. Enzyme-linked immunosorbant assay indicated that Cry1Ac protein accumulated at levels of 1-8 ng per gram of fresh tissue in leaves that were mechanically wounded. Transgenic sorghum plants were evaluated for resistance against the spotted stem borer (Chilo partellus Swinhoe) in insect bioassays, which indicated partial resistance to damage by the neonate larvae of the spotted stem borer. Reduction in leaf damage 5 days after infestation was up to 60%; larval mortality was 40%, with the surviving larvae showing a 36% reduction in weight over those fed on control plants. Despite the low levels of expression of Bt delta-endotoxin under the control of the wound-inducible promoter, the transgenic plants showed partial tolerance against first instar larvae of the spotted stem borer.

Genetic Transformation of Immature Sorghum Inflorescence via Microprojectile Bombardment

“…Shoot apices were bombarded and subcultured in a MS medium supplemented with benzylaminopurine (BAP) and naphthalene acetic acid (NAA). Leaf damage by the spotted stem borer (Chilo partellus) was reduced up to 60% in the sorghum transgenic plants generated (Girijashankar et al, 2005).…”

“…The biolistic has proved to be an efficient method for introducing new features in sorghum and a few transformation protocols are already available (Casas et al 1993;Casas et al 1997;Zhu et al 1998;Able et al 2001;Devi and Sticklen, 2002 ;Emani et al 2002;Tadesse et al, 2003;Girijashankar et al, 2005). The optimization of physical and biological parameters was the subject of most of the work published about sorghum transformation via bombardment.…”

“…Since the earliest 90's, laboratories around the world have generated improvements in sorghum regeneration and transformation that are ensuing in more consistent protocols. Transgenic sorghum plants have been generated via biolistic (Casas et al, 1993;Casas et al, 1997;Zhu et al, 1998;Able et al, 2001;Emani et al, 2002;Jeoung et al, 2002;Devi and Sticklen, 2003;Tadesse et al, 2003;Girijashankar et al, 2005) or Agrobacterium mediated transformation (Zhao et al, 2000;Carvalho et al, 2004;Gao et al, 2005;Howe et al, 2006;Van Nguyen et al, 2007;Nguyen et al, 2007;Gurel et al, 2009;Lu et al 2009;Mall et al, 2011).…”

Genetic transformation of Sorghum bicolor

“…Using the same promoter, Girijashankar et al, (2005) genetically transformed sorghum (genotype BTx 623) using particle bombardment method. Insect bioassays (leaf disc assay) of T 1 generation plants carrying mpicry1Ac showed reduction in leaf damage (60 %), larvae weight (25 %) and increased larval mortality (40 %) when compared to the control plants.…”

“…However, they did not confirm the integration or expression of the Bt gene in the putative transgenic plants. Girijashankar et al, (2005) also attempted to generate transgenic sorghum plants for resistance against the spotted stem borer (Chilo partellus Swinhoe). Shoot apices of sorghum genotype BTx 623 were transformed with synthetic ubi-cry1Ab, ubi-cry1Ac and mpiC1-cry1Ac using particle bombardment method (PIG).…”