Lectin and leghemoglobin in legumes play the important roles, respectively, in recognition of host plants to their rhizobial bacteria, and lowering the oxygen partial pressure around bacteroids and protecting nitrogen...Lectin and leghemoglobin in legumes play the important roles, respectively, in recognition of host plants to their rhizobial bacteria, and lowering the oxygen partial pressure around bacteroids and protecting nitrogenase from oxygen in symbiotic nitrogen-fixing nodules. In order to extend the host range of the rhizobial bacteria and to make them fix nitrogen in non-legumes, pea lectin gene (pl) and Parasponia hemoglobin gene ( phl,) have been constructed into a plant expression vector (pCBHUL) and the vector pCBHUL was introduced into rice calli from immature young embryos by particle bombardment. After the calli were regenerated into plantlets on the resistant-selecting media containing hygromycin, they were identified by PCR and Southern blot hybridization. It was indicated that the pi and phb genes were integrated into nucleic genome of the transformed rice plants. GUS activity and the product of the pi gene were determined by GUS staining, Western blot and in situ hybridization at translational level. Eighteen out of 40 plants resistant to hygromycin were positively identified by PCR analysis with the rate of 45%. The pi gene was expressed in 3 out of 18 plants with 17% and 7.5% in 40 plants. The results may provide a clue for exploring whether Rhizobium leguminosarum by. viceae could extend its host range and make the transgenic rice plants have the possibility of being symbiotic, or associative to nitrogen fixation.展开更多
Wheat, triticale, tritordeum, barley, oat and rye are the most important crops in human consumptions and industry in the world. Transformation technology supplies a new source of improving Triticeae crops. In the past...Wheat, triticale, tritordeum, barley, oat and rye are the most important crops in human consumptions and industry in the world. Transformation technology supplies a new source of improving Triticeae crops. In the past decade, transformation of wheat crops has considerably progressed. Many transgenic plants of Triticeae crops with various genes were produced via nricroprojectile bombardment, Agrobacterium-mediated transformation, PEG-uptake DNA technique, electroporation, microinjection, injection inflorescence and silicone carbide. Integration and expression of transgenes, inheritance and variation of transgenic plants have been studied. Technical improvements of genetic transformation for wheat crops will be extensively useful in commerce and benefit significantly to human being in the world.展开更多
Aim To induce and express the T4 lysozyme in Pichia pastoris and test the antibacterial activity of the protein. Methods T4 lysozyme gene was inserted into expression vector pPIC9K of Pichia pastoris with the fusion a...Aim To induce and express the T4 lysozyme in Pichia pastoris and test the antibacterial activity of the protein. Methods T4 lysozyme gene was inserted into expression vector pPIC9K of Pichia pastoris with the fusion at N terminal. The recombinant plasmid was digested by Sal I and then introduced into prepared GS115 competent cells by electroporation. Positive clone and multiple inserts were screened. The secreted proteins in the supernatants were tested. In the agar holes diffusion assay, our expressed protein showed significant antibacterial circles. Results T4 lysozyme protein inhibited the growth of staphylococcus aureus and streptococcus Pneumoniae. There was no difference in the bactericidal activity and the amount of protein expression between the single and multiple copies. The antibacterial activity of expressed protein remained the same during the heat stability test. Conclusion T4 lysozyme was successfully induced and expressed in Pichia pastoris. There is no relationship between copy number and expression. T4 lysozyme protein is heat stable.展开更多
Green fluorescent protein (GFP) gene was successfully transferred into the isolated zygotes and early proembryos of wheat (Triticum aestivum L.) by electroporation. A frequency, as high as 46.7% of GFP gene transient ...Green fluorescent protein (GFP) gene was successfully transferred into the isolated zygotes and early proembryos of wheat (Triticum aestivum L.) by electroporation. A frequency, as high as 46.7% of GFP gene transient expression in early proembryos, was achieved under 150 V/cm electric field strength, 25 muF capacitor, 200 mug/mL of linear plasmid DNA and an electroporation buffer at pH 7.2. Compared with five-day-old proembryos, the zygotes and early proembryos needed lower optimum strength of electric field. After culturing in KM8p medium, the electroporated early proembryos divided and GFP gene expression was observed in daughter cells and subsequent divisions. There was no mosaicism of gene expression in the zygotes and 2-, 4- and 8-celled proembryos.展开更多
文摘Lectin and leghemoglobin in legumes play the important roles, respectively, in recognition of host plants to their rhizobial bacteria, and lowering the oxygen partial pressure around bacteroids and protecting nitrogenase from oxygen in symbiotic nitrogen-fixing nodules. In order to extend the host range of the rhizobial bacteria and to make them fix nitrogen in non-legumes, pea lectin gene (pl) and Parasponia hemoglobin gene ( phl,) have been constructed into a plant expression vector (pCBHUL) and the vector pCBHUL was introduced into rice calli from immature young embryos by particle bombardment. After the calli were regenerated into plantlets on the resistant-selecting media containing hygromycin, they were identified by PCR and Southern blot hybridization. It was indicated that the pi and phb genes were integrated into nucleic genome of the transformed rice plants. GUS activity and the product of the pi gene were determined by GUS staining, Western blot and in situ hybridization at translational level. Eighteen out of 40 plants resistant to hygromycin were positively identified by PCR analysis with the rate of 45%. The pi gene was expressed in 3 out of 18 plants with 17% and 7.5% in 40 plants. The results may provide a clue for exploring whether Rhizobium leguminosarum by. viceae could extend its host range and make the transgenic rice plants have the possibility of being symbiotic, or associative to nitrogen fixation.
文摘Wheat, triticale, tritordeum, barley, oat and rye are the most important crops in human consumptions and industry in the world. Transformation technology supplies a new source of improving Triticeae crops. In the past decade, transformation of wheat crops has considerably progressed. Many transgenic plants of Triticeae crops with various genes were produced via nricroprojectile bombardment, Agrobacterium-mediated transformation, PEG-uptake DNA technique, electroporation, microinjection, injection inflorescence and silicone carbide. Integration and expression of transgenes, inheritance and variation of transgenic plants have been studied. Technical improvements of genetic transformation for wheat crops will be extensively useful in commerce and benefit significantly to human being in the world.
文摘Aim To induce and express the T4 lysozyme in Pichia pastoris and test the antibacterial activity of the protein. Methods T4 lysozyme gene was inserted into expression vector pPIC9K of Pichia pastoris with the fusion at N terminal. The recombinant plasmid was digested by Sal I and then introduced into prepared GS115 competent cells by electroporation. Positive clone and multiple inserts were screened. The secreted proteins in the supernatants were tested. In the agar holes diffusion assay, our expressed protein showed significant antibacterial circles. Results T4 lysozyme protein inhibited the growth of staphylococcus aureus and streptococcus Pneumoniae. There was no difference in the bactericidal activity and the amount of protein expression between the single and multiple copies. The antibacterial activity of expressed protein remained the same during the heat stability test. Conclusion T4 lysozyme was successfully induced and expressed in Pichia pastoris. There is no relationship between copy number and expression. T4 lysozyme protein is heat stable.
文摘Green fluorescent protein (GFP) gene was successfully transferred into the isolated zygotes and early proembryos of wheat (Triticum aestivum L.) by electroporation. A frequency, as high as 46.7% of GFP gene transient expression in early proembryos, was achieved under 150 V/cm electric field strength, 25 muF capacitor, 200 mug/mL of linear plasmid DNA and an electroporation buffer at pH 7.2. Compared with five-day-old proembryos, the zygotes and early proembryos needed lower optimum strength of electric field. After culturing in KM8p medium, the electroporated early proembryos divided and GFP gene expression was observed in daughter cells and subsequent divisions. There was no mosaicism of gene expression in the zygotes and 2-, 4- and 8-celled proembryos.
