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Development and evaluation of polyclonal antibody against wheat Rpl3
New Biotechnology · Volume 25S · September 2009
3.2.26 Isolation and activity of a protein cucumisin like of Solanum elaeagnifolium C.L. Vargas-Requena 1,∗ , F. Jimenez Vega 1 , A. Hernandez Santoyo 2 1
2
Universidad Autónoma de Ciudad Juarez, Ciudad Juarez, México, Mexico Universidad Nacional Autónoma de México, Mexico
Solanum elaeagnifolium is a plant has a negative impact mainly on crop production but also on livestock production, on the environment, and on trade and international relations. It is native to north-east Mexico and south-west USA where it is a weed. Other negative effects include hindering commercial cropping activities, harboring agricultural pests, poisoning livestock and reducing land values. The plant is officially declared as a noxious weed in several countries. In some parts of Chihuahua, México, the berry of this plant is used to make cheese in artisan form. In this research the protein with similar activity of rennet was obtained of an aqueous extract of the yellow berry of Solanum, because this is where the protease is found in greater quantity and activity. This extract was analyzed by ion exchange chromatography, isolated a protein cationic responsible for this activity. The molecular weight of this protein is approximately 19 kDa, their activity increases with increasing temperature, and remaining active even at 90 ◦ C. Its optimum pH of action is 5.2, in the pH of fresh milk, 6.8, has zero activity; by this reason in the actuality only is used to make asadero cheese. doi:10.1016/j.nbt.2009.06.659
3.2.27 Synonymous codon usage for expression enhancement in transgenic potato plants H. Ofoghi Biotechnology Department IROST, Tehran, Islamic Republic of Iran
It is well known that expression of foreign genes in genetically modified organisms can be low. This can result from an occurrence of sequence element similar to expression signals of a host located within the protein coding region. It is possible that transgene expression signals will be different from the signals for genes of a recipient organism. Despite the interrelationship between codon content and translation rate was proved for some bacteria, Saccharomyces cerevisiae and Drosophila, no strict evidence for such a phenomenon for higher plants was reported. However, it was frequently reported that an optimization of synonymous codon content could increase the transgene expression level. Genetic engineering of plants promises to create new opportunities in agriculture, environmental biology, production of chemicals and medicine. Due to high yields and functional maintenance at low expenses, biopharming and pharmaceuticals produced in plants is a rapidly expanding technology. However, expressing foreign genes in plants presents many technical
ABSTRACTS
challenges that are not encountered at this extent with other heterologous expression systems. It involves many aspects to be taken into account, some of which have been properly addressed in the past. Accordingly, cis-acting elements such as promoter, enhancer or terminator sequences have been studied carefully in order to optimize transgenic expression in plants and are readily available in convenient cloning cassettes. However, designing the actual coding region for each heterologous gene is very distinct and equally critical to achieve successful expression. In this study we synthesized a potato plant codon-optimized version of the human calcitonin coding sequence and introduced it into potato plant. To reveal the correlations between codon content and gene expression rate we have to analyze the expression level of recombinant hormone in transgenic potato. doi:10.1016/j.nbt.2009.06.660
3.2.28 Development and evaluation of polyclonal antibody against wheat Rpl3 F. Sanjarian ∗ , M. Mazaheri, A. Mousavi, B. Dehsara National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Islamic Republic of Iran
Trichothecenes, a large group of fusaritoxins, are potential inhibitors to eukaryotic protein via binding to Ribosomal Protein L3 (RPL3). The finding that trichothecene production contributes to virulence of Fusarium graminearum suggests that it may be possible to generate plants that are resistance to this fungus by increasing their resistance to them. Increase resistance against trichothecenes can result from over expression of RPL3 as well as modification of RPL3 in specific amino acid residues. In order to study the RPL3 expression profile in present and absent of toxin in Fusarium Head Blight susceptible and tolerant plants, specific antibodies are required. In this project we decided to produce polyclonal antibody against wheat RPL3. The over expressed wheat RPL3 antigen in prokaryotic host was purified from acryl amide gel and used for rabbit injection. Production of appropriate antibody was studied by using protein blot analysis, ELISA and Western blot analysis methods. doi:10.1016/j.nbt.2009.06.661
3.2.29 Phytoremediation of anthracene by transgenic tobacco plants with a fungal glutathione-S-transferase gene P. Dixit ∗ , P. Mukherjee, P. Sherkhane, S. Kale, S. Eapen Bhabha Atomic Research Centre, Mumbai, India
Polycyclic aromatic hydrocarbons (PAHs) such as anthracene and pyrene are pollutants produced via natural and anthropogenic sources, generated during the incomplete combustion of solid and liquid fuels or derived from industrial activities and are known to
www.elsevier.com/locate/nbt S291
3.2.26 Isolation and activity of a protein cucumisin like of Solanum elaeagnifolium C.L. Vargas-Requena 1,∗ , F. Jimenez Vega 1 , A. Hernandez Santoyo 2 1
2
Universidad Autónoma de Ciudad Juarez, Ciudad Juarez, México, Mexico Universidad Nacional Autónoma de México, Mexico
Solanum elaeagnifolium is a plant has a negative impact mainly on crop production but also on livestock production, on the environment, and on trade and international relations. It is native to north-east Mexico and south-west USA where it is a weed. Other negative effects include hindering commercial cropping activities, harboring agricultural pests, poisoning livestock and reducing land values. The plant is officially declared as a noxious weed in several countries. In some parts of Chihuahua, México, the berry of this plant is used to make cheese in artisan form. In this research the protein with similar activity of rennet was obtained of an aqueous extract of the yellow berry of Solanum, because this is where the protease is found in greater quantity and activity. This extract was analyzed by ion exchange chromatography, isolated a protein cationic responsible for this activity. The molecular weight of this protein is approximately 19 kDa, their activity increases with increasing temperature, and remaining active even at 90 ◦ C. Its optimum pH of action is 5.2, in the pH of fresh milk, 6.8, has zero activity; by this reason in the actuality only is used to make asadero cheese. doi:10.1016/j.nbt.2009.06.659
3.2.27 Synonymous codon usage for expression enhancement in transgenic potato plants H. Ofoghi Biotechnology Department IROST, Tehran, Islamic Republic of Iran
It is well known that expression of foreign genes in genetically modified organisms can be low. This can result from an occurrence of sequence element similar to expression signals of a host located within the protein coding region. It is possible that transgene expression signals will be different from the signals for genes of a recipient organism. Despite the interrelationship between codon content and translation rate was proved for some bacteria, Saccharomyces cerevisiae and Drosophila, no strict evidence for such a phenomenon for higher plants was reported. However, it was frequently reported that an optimization of synonymous codon content could increase the transgene expression level. Genetic engineering of plants promises to create new opportunities in agriculture, environmental biology, production of chemicals and medicine. Due to high yields and functional maintenance at low expenses, biopharming and pharmaceuticals produced in plants is a rapidly expanding technology. However, expressing foreign genes in plants presents many technical
ABSTRACTS
challenges that are not encountered at this extent with other heterologous expression systems. It involves many aspects to be taken into account, some of which have been properly addressed in the past. Accordingly, cis-acting elements such as promoter, enhancer or terminator sequences have been studied carefully in order to optimize transgenic expression in plants and are readily available in convenient cloning cassettes. However, designing the actual coding region for each heterologous gene is very distinct and equally critical to achieve successful expression. In this study we synthesized a potato plant codon-optimized version of the human calcitonin coding sequence and introduced it into potato plant. To reveal the correlations between codon content and gene expression rate we have to analyze the expression level of recombinant hormone in transgenic potato. doi:10.1016/j.nbt.2009.06.660
3.2.28 Development and evaluation of polyclonal antibody against wheat Rpl3 F. Sanjarian ∗ , M. Mazaheri, A. Mousavi, B. Dehsara National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Islamic Republic of Iran
Trichothecenes, a large group of fusaritoxins, are potential inhibitors to eukaryotic protein via binding to Ribosomal Protein L3 (RPL3). The finding that trichothecene production contributes to virulence of Fusarium graminearum suggests that it may be possible to generate plants that are resistance to this fungus by increasing their resistance to them. Increase resistance against trichothecenes can result from over expression of RPL3 as well as modification of RPL3 in specific amino acid residues. In order to study the RPL3 expression profile in present and absent of toxin in Fusarium Head Blight susceptible and tolerant plants, specific antibodies are required. In this project we decided to produce polyclonal antibody against wheat RPL3. The over expressed wheat RPL3 antigen in prokaryotic host was purified from acryl amide gel and used for rabbit injection. Production of appropriate antibody was studied by using protein blot analysis, ELISA and Western blot analysis methods. doi:10.1016/j.nbt.2009.06.661
3.2.29 Phytoremediation of anthracene by transgenic tobacco plants with a fungal glutathione-S-transferase gene P. Dixit ∗ , P. Mukherjee, P. Sherkhane, S. Kale, S. Eapen Bhabha Atomic Research Centre, Mumbai, India
Polycyclic aromatic hydrocarbons (PAHs) such as anthracene and pyrene are pollutants produced via natural and anthropogenic sources, generated during the incomplete combustion of solid and liquid fuels or derived from industrial activities and are known to
www.elsevier.com/locate/nbt S291