Konstruksi Vektor Biner Mengandung Gen Hibrid cryIB-cryIAa untuk Transformasi Agrobacterium Tanaman Padi (Oryza sativa L)

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

Studi Agronomis Tanaman Padi (Oryza sativa L.) Hasil Ko-Kultivasi Beberapa Strain Agrobacterium tumefaciens

The transgenic rice of Cisadane and Rajalele cultivars (cv) was obtained by co-cultivation of rice callus by 4 strain Agrobacterium tumefaciens containing 4 different binary vector (pIG 121 Hm, pCAMBIA 1301, pTOK 233 and pMOG22 BarGus). The transgenic plantlets were acclimatized and grown in the growth chamber to investigate their agronomic characteristic in comparison to control plants. The results showed that there was no difference in the number of spikelet and seeds/spikelet among different transgenic lines co-cultivated by different strains. The number of seed/spikelet of transgenic rice cv. Rojolele and Cisadane compared to their control did not show significant difference. However, the total number of spikelet was different only for cv. Rojolele

THE SEGREGATION PATTERN OF INSECT RESISTANCE GENES IN THE PROGENIES AND CROSSES OF TRANSGENIC ROJOLELE RICE

Successful application of genetic transformation technique, especially in developing rice variety resistant to brown plant hopper and stem borer, will depend on transgene being expressed and the gene inherited in a stable and predictable manner. This study aimed to analyse transgene segregation pattern of the progenies and the crosses of transgenic rice cv. Rojolele harboring cry1Ab and gna genes. The third generation (T2) of fivetransgenic Rojolele events containing gna and/or cry1Ab were evaluated for two generations to identify the homozygous lines and to study their inheritance. The homozygous lines were selected based on the result of PCR technique. The segregation patterns of gna and cry1Ab were studied in eight F2 populations derived from Rojolele x transgenic Rojolele homozygous for cry1Ab and or gna and their reciprocal crosses. Data  resulted from PCR of F2 population were analysed using a Chi Square test. The study obtained six homozygous lines for gna, namely A22- 1-32, A22-1-37, C72-1-9, F11-1-48, K21-1-39, K21-1-48, and two homozygous lines for cry1Ab, namely K21-1-39 and K21- 1-48. Both cry1Ab and gna transgenes had been inherited through selfing and crossing with their wild type as indicated from the F1 containing gna and cry1Ab as many as 48.4% and 47.4%, respectively. In six of the eight crosses, gna was inherited in a 3:1 ratio consistent with Mendelian inheritance of a single dominant locus, while in the remaining two crosses, gna was segregated in a 1:1 ratio. The presence of cry1Ab in F2 populations also showed a 3:1 segregation ratio in all crosses. In the F2 population derived from F1 plant containing cry1Ab and gna, both transgenes segregated in a 9:3:3:1 dihybrid segregation ratio. This study will add to the diversity of genetic sources for insect resistance and allow further use of these transgenic lines for pyramiding resistance to brown plant hopper and stem borer or  separately in rice breeding programs whenever the efficacy tests and biosafety requirements have been completed

Keefektivan Padi Transgenik terhadap Hama Penggerek Batang Padi Kuning Scirpophaga incertulas (Walker) (Lepidoptera: Crambidae).

Transformation two cry genes (cryIB-cryIAa) and transformation with a single the cry1B gene under the control of a wound-inducible maize proteinase inhibitor gene (mpi) promoter were two approaches that were used to get resistant rice to the rice stemborer which may be had a durable resistance. To obtain information on the effectiveness of seven transgenic rice lines to the rice yellow stemborer S. incertulas, a test was conducted in greenhouse. The seven lines were 1). line 4.2.3 and 2) line 4.2.4 both contain fusion of two cry genes (cryIB-cryIAa); 3) line 3R9 and 4) line 3R7 lines both contain of mpi-cryIB gene; and 5) line 6.11 contains of cryIAb gene by particle bombardment, 6) line DT-cry (Azygous) that do not contain cry gene (null), and 7) DT-cry line contains cryIAb gene by Agrobacterium, and as a negative control, we used three non transgenic rice varieties i.e., Rojolele, Cilosari, and Ciherang. The result showed that transgenic rice lines, except DT-cry and DT-cry (Azygous) lines were effective to suppress damage by the insect, and showed an inhibition effect on the growth of S. incertulas, and had a high level ofresistance than non transgenic rice varieties had. There were differences on resistance value/level among transgenic rice lines. Based on the resistance value, 6.11 line was the highest (scale 0) followed by 4.2.4 line and 3R7 line, these lines were categorized as high resistance (scale 1). Transgenic rice-4.2.3 line and 3R9 line were categorized as moderat resistance (scale 3). DT-cry and DT-cry (Azygous) lines were susceptible (scale 7-9). KEY WORDS: Effectiveness, resistance value, transgenic rice, S. incertulas

Keefektivan Padi Transgenik terhadap Hama Penggerek Batang Padi Kuning Scirpophaga incertulas (Walker) (Lepidoptera: Crambidae).

Transformation two cry genes (cryIB-cryIAa) and transformation with a single the cry1B gene under the control of a wound-inducible maize proteinase inhibitor gene (mpi) promoter were two approaches that were used to get resistant rice to the rice stemborer which may be had a durable resistance. To obtain information on the effectiveness of seven transgenic rice lines to the rice yellow stemborer S. incertulas, a test was conducted in greenhouse. The seven lines were 1). line 4.2.3 and 2) line 4.2.4 both contain fusion of two cry genes (cryIB-cryIAa); 3) line 3R9 and 4) line 3R7 lines both contain of mpi-cryIB gene; and 5) line 6.11 contains of cryIAb gene by particle bombardment, 6) line DT-cry (Azygous) that do not contain cry gene (null), and 7) DT-cry line contains cryIAb gene by Agrobacterium, and as a negative control, we used three non transgenic rice varieties i.e., Rojolele, Cilosari, and Ciherang. The result showed that transgenic rice lines, except DT-cry and DT-cry (Azygous) lines were effective to suppress damage by the insect, and showed an inhibition effect on the growth of S. incertulas, and had a high level ofresistance than non transgenic rice varieties had. There were differences on resistance value/level among transgenic rice lines. Based on the resistance value, 6.11 line was the highest (scale 0) followed by 4.2.4 line and 3R7 line, these lines were categorized as high resistance (scale 1). Transgenic rice-4.2.3 line and 3R9 line were categorized as moderat resistance (scale 3). DT-cry and DT-cry (Azygous) lines were susceptible (scale 7-9). KEY WORDS: Effectiveness, resistance value, transgenic rice, S. incertulas

Suplement Agricultural Biotechnology Principle and Practice : A Compilation of Lecture Presentations of Speakers from japan and Philippines

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