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All Journal ANNALES BOGORIENSES Annales Bogorienses
Estiati, Amy
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Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Chemical Engineering, Chemistry & Bioengineering Chemistry Immunology & microbiology Medicine & Pharmacology

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Overexpression of OsHox-6 Gene Enhanced Tiller Number in Rice But Induced Yield Penalty Rahmawati, Syamsidah; Chairunnisa, Chairunnisa; Erdayani, Eva; Nugroho, Satya; Estiati, Amy
ANNALES BOGORIENSES Vol 23, No 1 (2019): Annales Bogorienses
Publisher : Research Center for Biotechnology - Indonesian Institute of Sciences (LIPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (712.857 KB) | DOI: 10.14203/ann.bogor.2019.v23.n1.30-40

Abstract

OsHox-6, belongs to the transcription factor homeodomain leucine zipper (HD-Zip) protein sub-family I, has unknown function. This study was aimed to characterize the phenotypes of two homozygous transgenic rice lines (S29-62-2 and S.40.4-158-1) containing an extra copy of OsHox-6 gene under the control of a rice constitutive promoter, OsLEA3, and to evaluate their tolerance to water stress. A real-time quantitative PCR (qRT-PCR) showed that the transcript expression of OsHox-6 gene in the transgenic lines increased 5-10 folds under a normal irrigation and 10-20 folds after exposure to water stress conditions as compared to its wild type control. Transgenic plants overexpressing OsHox-6 exhibited phenotypic alteration at the normal irrigation by inducing tiller formation, suggesting a decrease in the apical dominance. Transgenic plants also showed significant enhancement in the total grain number, however, the number of empty grains  also increased significantly (~16-22%).  After imposed to the water stress, the number of empty grains in the transgenic lines was even higher (up to 83% in average). Furthermore, observations on the water loss rates, relative water contents and drought resistance indices (DRI) suggested that the overexpression of OsHox-6 did not significantly increase tolerance to water stress.  Further research is required to reveal the detailed mechanisms of OsHox-6 in response to water and other abiotic stresses.
Overexpression of OsHox-6 Gene Enhanced Tiller Number in Rice But Induced Yield Penalty Rahmawati, Syamsidah; Chairunnisa, Chairunnisa; Erdayani, Eva; Nugroho, Satya; Estiati, Amy
Annales Bogorienses Vol. 23 No. 1 (2019): Annales Bogorienses
Publisher : BRIN

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

OsHox-6, belongs to the transcription factor homeodomain leucine zipper (HD-Zip) protein sub-family I, has unknown function. This study was aimed to characterize the phenotypes of two homozygous transgenic rice lines (S29-62-2 and S.40.4-158-1) containing an extra copy of OsHox-6 gene under the control of a rice constitutive promoter, OsLEA3, and to evaluate their tolerance to water stress. A real-time quantitative PCR (qRT-PCR) showed that the transcript expression of OsHox-6 gene in the transgenic lines increased 5-10 folds under a normal irrigation and 10-20 folds after exposure to water stress conditions as compared to its wild type control. Transgenic plants overexpressing OsHox-6 exhibited phenotypic alteration at the normal irrigation by inducing tiller formation, suggesting a decrease in the apical dominance. Transgenic plants also showed significant enhancement in the total grain number, however, the number of empty grains also increased significantly (~16-22%). After imposed to the water stress, the number of empty grains in the transgenic lines was even higher (up to 83% in average). Furthermore, observations on the water loss rates, relative water contents and drought resistance indices (DRI) suggested that the overexpression of OsHox-6 did not significantly increase tolerance to water stress. Further research is required to reveal the detailed mechanisms of OsHox-6 in response to water and other abiotic stresses.
Agrobacterium-Mediated Transformation of Javanica Rice Plants With A Cry1b Gene Under The Control of Wound-Inducible Gene Promoter Estiati, Amy; Rachmawati, Syamsidah; Astuti, Dwi; Loedin, Inez Hortenza Slamet
Annales Bogorienses Vol. 11 No. 1 (2007): Annales Bogorienses
Publisher : BRIN

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Abstract

A cry1B synthetic gene of Bacillus thuringiensis has been used for the transformation of the javanica rice plants cv. Rojolele to confer resistance to an important pest yellow stem borer (Scirpophaga incertulas). Embryogenic callus were co-cultivated with the EHA105 strain of Agrobacterium tumefaciens harbouring binary vector pCAMBIA I301 containing cry1B gene under the control of wound inducible gene promoter (mpi), hygromycin resistance gene (hpt) as a selectable marker and intron-containing b-glucuronidase (gus intron) gene as a reporter gene driven by CaMV35S promoter. Previously. our histochemical assay and PCR analysis had proved the integration of cry1B gene into the genome of rice plants at first generation. However, the existence of the gene should remain stable throughout generation. In this study, the presence of the cry1B transgene in rice transgenic plants at second generation was confirmed by Polymerase Chain Reaction (PCR). Insertion of the cry1B gene in the genome of PCR positive plants was verified by Southern blot analysis and showed that integration of cry1B into the genomic DNA of javanica rice plants cv. Rojolele. An effective resistance of transgenic plants against stem borer was verified in bioassays.
Agrobacterium-mediated Transformation of Banana Musa acuminata AA cv "“Mas Lampung” with hpt Gene Using Sterile Corm as Target Tissue Estiati, Amy; Nena, Ade; Witjaksono, Witjaksono
Annales Bogorienses Vol. 13 No. 1 (2009): Annales Bogorienses
Publisher : BRIN

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Abstract

The protocol for Agrobacterium-mediated transformation of local banana plants cv “Mas Lampung” (AA) has been established. A selectable marker gene (hpt) has been used to study the transformation using in vitro corm slices as target tissues. Banana in vitro corm slices were co-cultivated with the EHA105 strain of Agrobacterium tumefaciens harbouring binary vector pCAMBIA 1301 containing hygromycin resistance gene (hpt) as a selectable marker and intron-containing β-Glucuronidase (gus-intron) gene as a reporter gene driven by CaMV 35S promoter. Polymerase Chain Reaction (PCR) were used to examine the existence of hpt gene in plants resulted from the transformation. Using primer pairs specific for hpt gene, our PCR analysis on leaves showed the presence of the hpt transgene in banana transgenic plants at first generation (T0) of transformation. To prove the existence of hpt gene in the fruits of transgenic banana plants, PCR analysis were also carried out. The data showed that the hpt gene could be amplified from banana fruits of tested samples. These result demonstrates that the Agrobacterium-mediated transformation method used in this experiment has been successful to transfer gene into banana plants. Thus, the transformation method reported here could be used as a standard protocol to transfer another useful genes into local banana plants cv. “Mas Lampung”. Furthermore, the presence of transgene in fruits of banana transgenic plants is important achievement especially for transgene that is expected to be expressed in the fruit including to introduce vaccine genes into banana fruits for edible vaccine.
Gene Flow from Genetically Modified Rice to Their Weedy and Wild Relatives and Its Environmental Consequences Estiati, Amy; Nugroho, Satya
Annales Bogorienses Vol. 13 No. 1 (2009): Annales Bogorienses
Publisher : BRIN

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Abstract

Rice is one of the most important food crops in the world. Nearly half of the world population consume rice as their staple food (FAO, 2004). With the increasing of world population, the need to provide more food supplies including rice is obvious. Biotechnology is expected to play major roles in the improvement of crop productivity and quality. Unlike transgenic maize, eventhough many research have been done to improve rice performance by genetic engineering, transgenic rice has not been released or commercialized. Among those research to improve rice performance are the attempt to introduce biotic and abiotic stress tolerant traits, herbicide tolerant trait and beta-carotin biosynthetic pathway. Currently, several genetically modified rice cultivars have been and being tested on limited field trials. Undoubtedly, biotechnology will benefit agriculture and thus providing enough food source to keep up with the ever-increasing needs in the future. However, the release and utilization of such technology in agriculture is still arising concerns about the impact to the environment. Therefore the possibility of the transgene escape to the environment needs to be analyzed. For example, whether the application of such technology can create superweed that resulted in the environment problems. On the other hand, cultivated rice may acquire genes for weediness from pollen-mediated gene flow of weedy or wild rice occurring inside or near cultivated rice fields, which leads to persistence and invasiveness of the cultivated rice, although the chance is low. This article discuss the possibility of the occurrence gene flow from the application of genetically modified rice to surrounding rice plants including its wild relatives and weed, and the requirements or precautions needed to be done to prevent gene flow. 
Segregation Analysis of Transgenic Rice Plants cv Rojolele Harboring cry1B Gene and Plant Selection for Potential to Yellow Stem Borer Estiati, Amy; Sulistyowati, Yuli; Zahra, Fatimah; Nurhasanah, Ade Nena; Rachmawati, Syamsidah; Loedin, Inez Hortenza Slamet
Annales Bogorienses Vol. 16 No. 1 (2012): Annales Bogorienses
Publisher : BRIN

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Abstract

Transgenic rice plants harboring resistant gene to yellow stem borer, cry1B gene had been obtained. However, the cry1B gene in second generation of transgenic rice lines still segregating following Mendelian ratio 3:1. For further use in the breeding programs, it is important to ensure that the gene is dominant gene and the plants are homozygous for cry1B gene. Selection of homozygous transgenic rice lines containing cry1B gene at third, fourth and fifth generation had been conducted by PCR. The presence of cry1B gene was determined by showing the PCR product of 1.9 kb. Segregation analysis proved that six transgenic rice lines i.e. 3R7.8.15.1, 3R7.8.15.9, 3R7.8.15.10, 3R7.8.15.15, 3R7.8.15.21, 3R7.8.15.29 are homozygous lines for cry1B gene. Moreover, bioassay studies at vegetative stage on six homozygous transgenic rice lines showed that these transgenic rice lines are potential resistant to yellow stem borer comparing with non-transgenic plants, with the score of 0 (indicated no symptom) for six transgenic lines and score of 9 (more than 60% damaged tillers ) for non-transgenic plants. However, to confirm the efficacy of cry1B gene to yellow stem borer in natural condition, confined field trial in endemic area of yellow stem borer should be conducted. 
Insect Bioassay in Biosafety Containment to Select Transgenic Rice (Oryza sativa L.) Harboring Cry1B Gene Resistant to Yellow Stem Borer (Scrirpophaga incertulas Walk.) Estiati, Amy; Nurhasanah, Ade Nena; Nugroho, Satya
Annales Bogorienses Vol. 17 No. 2 (2013): Annales Bogorienses
Publisher : BRIN

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Abstract

Development of rice varieties resistant to yellow stem borer (YSB) is very crucial. Agrobacterium-mediated transformation of cry1B gene under wound inducible gene promoter mpi (maize proteinase inhibitor) into a local rice variety Rojolele had been conducted. PCR analysis proved that cry1B gene had been integrated into plant genome of 3R25 and 3R5 rice lines. Segregation analysis using PCR for cry1B gene of the two putative transgenic rice lines at third (T2), fourth (T3), fifth (T4), and sixth (T5) generations of 3R25 and 3R5 lines proved that 3R25.7.27, 3R25.7.13.8.2, 3R25.7.13.8.6, 3R25.7.13.8.8, 3R5.26.2, and 3R5.26.5 are homozygous lines for cry1B gene. Insect bioassay on three randomly picked homozygous transgenic rice lines to study the efficacy of cry1B gene toward YSB was conducted in biosafety containment by infestingYSB larvae at first instar into 3R25.7.27, 3R25.7.13.8.6, and 3R5.26.5 transgenic rice lines, using non--transgenic Rojolele, IR64 and IR74 as susceptible controls. The results showed that the percentages of deadhearts symptoms of 3R25.7.27, 3R5.26.2, and 3R25.7.13.8.6 rice lines were lower than those of the susceptible control lines with scores of 0.1 and 0, respectively. While the scores of all three susceptible control plants were 9. The results proved that lines 3R25.7.27, 3R25.7.13.8.6, and 3R5.26.2 were categorized as resistant lines while the non-transgenic Rojolele, IR64, and IR74 were categorized as susceptible lines. The results also showed that the cry1B gene was expressed and produced insecticidal protein CRY1B which were active against YSB to protect rice plant toward YSB infestation.
Construction of Binary Vector With Wound Inducible Promoter for HbsAg Expression: Development of Plant-Based Edible Hepatitis B Vaccine from Indonesian Isolate Suhandono, Sony; Giri-Rachman, Ernawati A.; Zainuddin, Ima M.; Utari, Putri Dwi; Supraba, Apsari; Estiati, Amy
Annales Bogorienses Vol. 11 No. 1 (2007): Annales Bogorienses
Publisher : BRIN

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Abstract

Hepatitis B is a serious infectious disease in the third world countries including Indonesia. Vaccination is the most effective way to prevent the spread of the disease; therefore the demand for HBV vaccine is high. In order to produce more vaccine at lower cost, transgenic plant can be chosen to express the vaccine with the above criteria. Several researches were successfully producing transgenic plants expressing HBsAg that formed virus-like particles and induced immune response in human. However, HBsAg expression in transgenic plant needs to be improved especially on gene expression control system. Here, we describe the construction of HBsAg structural gene under the control of wound inducible promoter, MeEF1 promoter from Manihot esculenta Crantz. The HBsAg gene was amplified using PCR from HBV genome isolated from an Indonesian patient. The gene was subsequently fused with VSPaS signal peptide, which targeted the reticulum endoplasm of plant cell. The construct was cloned into binary expression vector for Agrobacterium plant transformation in near future.
Co-Authors Chairunnisa Chairunnisa Dwi Astuti Erdayani, Eva Ernawati A. Giri-Rachman Fatimah Zahra Loedin, Inez Hortenza Slamet Nena, Ade Nurhasanah, Ade Nena Rachmawati, Syamsidah Satya Nugroho SONY SUHANDONO Supraba, Apsari Syamsidah Rahmawati, Syamsidah Utari, Putri Dwi Witjaksono Witjaksono Yuli Sulistyowati Zainuddin, Ima M.