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All Journal Biota: Jurnal Ilmiah Ilmu-Ilmu Hayati Indonesian Journal of Biotechnology
Syamsidah Rahmawati
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Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Immunology & microbiology Materials Science & Nanotechnology

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Comparative Analysis of Rice Transformation Using Agrobacterium tumefaciens and Rhyzobium leguminosarum Syamsidah Rahmawati; Osmat Azzam Jefferson; Didy Sopandie; S. Suharsono; Inez Hortense Slamet-Loedin
Indonesian Journal of Biotechnology Vol 15, No 1 (2010)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (287.394 KB) | DOI: 10.22146/ijbiotech.7821

Abstract

This study was aimed to study the effectiveness of Rhizobium transformation system compared to the most widely used Agrobacterium mediated transformation system on three rice cultivars, Ciherang (Indica), Nipponbare (Japonica), and Rojolele (Javanica). Six day old calli induced from immature embryos were inoculated with Rhizobium leguminosarum bv trifolii ANU845 and Agrobacterium tumefaciens LBA288 that harbored with vector pCAMBIA 5106. This plasmid contained a minimum set of transfer machinery genes and had a gusplus and an hptII gene driven by 35S CaMV promoter in the T-DNA. The results showed that the transformation frequencies (number of PCR positive plants per number of calli inoculated) ranging from 0 to 12.05 % depend on the genotype and transfer agent used. The highest transformation frequency (12.05%) was obtained in Ciherang transformed with R. leguminosarum. Most of the transgenic rice obtainedby Rhizobium transformation were normal in morphology and fertile similar to those obtained by Agrobacterium transformation. Integration, expression and inheritance of transgenes were demonstrated by molecular and genetic analysis in T0 and T1 generations.Key words : Rhizobium leguminosarum, immature embryos, Agrobacterium tumefaciens
Konstruksi Vektor Biner Mengandung Gen Hibrid cryIB-cryIAa untuk Transformasi Agrobacterium Tanaman Padi (Oryza sativa L) Syamsidah Rahmawati; Inez H. Slamet-Loedin
Biota : Jurnal Ilmiah Ilmu-Ilmu Hayati Vol 9, No 2 (2004): June 2004
Publisher : Universitas Atma Jaya Yogyakarta

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24002/biota.v9i2.2892

Abstract

This study aimed to obtain binary vectors carrying  cryIB-cryIAa hybrid gene suitable for Agrobacterium-mediated transformation of two Indonesian rice cultivars, Rajalele (Javanica) and Cisadane (Indica).  Introduction of hybrid Bt toxin, encoded by cryIB-cryIAa hybrid gene with different binding receptor in the insect gut driven by maize ubiquitin promoter, was intended to reduce the possibility of the appearance of stem borer resistant to Bt toxin. A HindIII fragment containing the ubi cryIB-cryIAa hybrid gene was excised from pBKS ubi cryIB-cryIAa and cloned into the HindIII site of binary vector pCAMBIA 1301, 1303, or 1304.  Restriction analysis of recombinant plasmids confirmed the insertion of the gene and demonstrated that the gene inserted in two orientations. The recombinant plasmids were transformed into electrocompetent Agrobacterium tumefaciens EHA105 by electroporation.  Embryogenic calli of Rajalele and Cisadane were co-cultivated with A. tumefaciens EHA 105 harbouring binary vector pCAMBIA 1301, 1303, or 1304 containing ubi cryIB-cryIAa. GUS histochemical assay revealed that high transformation efficiency (75-100%) on both cultivars were obtained from embryogenic calli transformed with pCAMBIA 1303 or 1304.
Co-Authors Didy Sopandie INEZ H. SLAMET-LOEDIN Inez Hortense Slamet-Loedin Osmat Azzam Jefferson S. Suharsono