Agrobacterium-mediated co-transformation of rice using two selectable marker genes derived from rice genome components
- First Online:
- 1.3k Downloads
A method for Agrobacterium-mediated co-transformation of rice (Oryza sativa L.) was developed using rice-derived selection markers. Two T-DNAs were efficiently introduced into separate loci using selectable marker gene cassettes consisting of the mutated acetolactate synthase gene (mALS) under the control of the callus-specific promoter (CSP) (CSP:mALS) and the ferredoxin nitrite reductase gene (NiR) under the control of its own promoter (NiR P:NiR). The CSP:mALS gene cassette confers sulfonylurea herbicide resistance to transgenic rice callus. The NiR P:NiR construct complements NiR-deficient mutant cultivars such as ‘Koshihikari’, which are defective in the regulation of nitrogen metabolism. In the present study, the CaMV35S:GUS and CaMV35S:GFP gene cassettes were co-introduced into the ‘Koshihikari’ genome using our system. Approximately 5–10 independent transgenic lines expressing both the GUS and GFP reporters were obtained from 100 Agrobacterium co-inoculated calli. Furthermore, transgenic ‘Koshihikari’ rice lines with reduced content of two major seed allergen proteins, the 33 and 14–16 kDa allergens, were generated by this co-transformation system. The present results indicate that the generation of selectable antibiotic resistance marker gene-free transgenic rice is possible using our rice-derived selection marker co-transformation system.
Key message An improved rice transformation method was developed based on Agrobacterium-mediated co-transformation using two rice genome-derived selectable marker gene cassettes.
KeywordsAcetolactate synthase Agrobacterium Co-transformation Ferredoxin nitrite reductase GM crop Transgenic rice
Green fluorescent protein
- GM crop
Genetically modified crop
Mutated acetolactate synthase
Ferredoxin nitrite reductase
- Baisakh N, Rehana S, Rai M, Oliva N, Tan J, Maclill D, Khush GS, Datta K, Datta SK (2006) Marker-free transgenic (MFT) near-isogenic introgression lines (NILs) of ‘golden’ indica rice (cv IR64) with accumulation of provitamin A in the endosperm tissue. Plant Biotechnol J 4:467–475PubMedCrossRefGoogle Scholar
- Ito S, Rosegrant MW, Agcaoili-Sombilla MC (1995) Quality-equivalent and cost-adjusted measurement of international competitiveness in Japanese rice markets. International Food Policy Research Institute, EPTD discussion paper no. 91, pp 1–23Google Scholar
- James C (2011) Global status of commercialized biotech/GM crops: 2011. ISAAA Brief No. 43, ISAAA, Ithaca, NYGoogle Scholar
- Matsuda T, Nakasa M, Alvarez AM, Izumi H, Kato T, Tada Y (2006) Rice-seed allergenic protein and hypoallergenic rice. In: Mine Y, Shahidi F (eds) Nutraceutical proteins and peptides in health and disease. Taylor and Francis, London, pp 493–511Google Scholar