In vitro culture and genetic transformation of sorghum by microprojectile bombardment

ABSTRACT

The pBluescript construct pSS1, with chitinase gene, isolated from American elm, and bar gene was transferred to sorghum by particle bombardment. The construct, containing the 1.2-kb chitinase gene (ECH2), driven by the Act promoter and Pin terminator, and the bar selectable marker gene driven by the CaMV 35 S promoter and nos terminator, was coated on tungsten particles which were used to bombard sorghum tissue. The target tissue, comprising multiple-shoot clumps, was obtained from the shoot apical meristems of in vitro-grown seedlings. The bombarded tissue was cultured on shoot multiplication medium containing the selective agent bialophos (3–5 mg / 1). Eleven putative T0 transgenic plants regenerated on selection medium were transferred to sterile soil mix and maintained in the containment glasshouse. The presence of chitinase and bar genes in five T0 transgenic plants was confirmed by PCR and Southern blot analysis. Studies on the inheritance pattern of herbicide resistance and fungal resistance are in progress.

KEY WORDS:

• Sorghum
• microprojectile bombardment
• transformation
• herbicide tolerance
• chitinase
• fungal disease resistance