title: Agrobacterium-mediated transformation of leaf base derived callus tissues of popular indica rice (Oryza sativa L. sub sp. indica cv. ADT 43).
abstract: A simple and efficient protocol for the Agrobacterium-mediated transformation of an agronomically useful abiotic sensitive popular indica rice cv. ADT 43 has been developed. Initiation of calli were best achieved from the leaf bases of 4 days old rice seedlings on LS medium supplemented with 2.5mg/L 2,4-D and 1.0mg/L thiamine-HCl. Rice calli immersed in Agrobacterium suspension (strain EHA 105, OD(600)=0.8) were co-cultured on LS30-AsPC medium for 2 days at 25±2°C in the dark. Based on GUS expression analysis, 10min co-cultivation time with 100μM acetosyringone was found optimum for the delivery of gus gene. Calli were proved to be very sensitive to Agrobacterium infection and we found that the level of necrotic response can be minimized after co-cultivation with 30% LS, 10g/L PVP, 10% coconut water and 250mg/L timentin which improved the final transformation efficiency to 9.33%. Molecular and genetic analysis of transgenic plants reveals the integration, expression and inheritance of transgene in the progeny (T(1)) of these plants. The copy number of transgenes has been found to vary from 1 to 2 in transgenic plants (T(0) and T(1)).
authors.affiliation: Department of Biotechnology, Alagappa University, Karaikudi, Tamil Nadu 630003, India.
authors.forename: Alagarsamy
authors.initials: A
authors.lastname: Karthikeyan
authors.forename: Shunmugiah Karutha
authors.initials: SK
authors.lastname: Pandian
authors.forename: Manikandan
authors.initials: M
authors.lastname: Ramesh
mesh_terms: D000098:Acetophenones / Q000008:administration & dosage; D018929:Cell Culture Techniques / Q000379:methods; D003470:Culture Media / Q000737:chemistry; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D012275:Oryza / Q000235:genetics* / Q000254:growth & development / Q000382:microbiology*; D010937:Plant Growth Regulators / Q000378:metabolism; D018515:Plant Leaves / Q000235:genetics / Q000254:growth & development / Q000382:microbiology; D030821:Plants, Genetically Modified / Q000235:genetics / Q000254:growth & development / Q000382:microbiology; D012231:Rhizobium / Q000235:genetics*; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D000098:Acetophenones; D003470:Culture Media; D010937:Plant Growth Regulators; C051667:acetosyringone