13 results about "Yellow fluorescent protein" patented technology

The invention discloses application of a PUB62 gene in regulating and controlling a plant to resist geminivirus and a transgenic plant cultivation method. The geminivirus is soybean yellow leaf curl virus, tomato yellow leaf curl virus and Yunnan tomato leaf curl virus. A nicotiana benthamiana PUB62 gene is cloned, the nicotiana benthamiana PUB62 gene is constructed into an NbPUB62-YFP vector with yellow fluorescent protein, the NbPUB62-YFP vector is introduced into plant cells through an agrobacterium-mediated transformation method, and an NbPUB62 overexpression transgenic plant is obtained through a cell and tissue culture technology and can remarkably inhibit infection of the soybean yellow leaf curl virus; meanwhile, the infection of tomato yellow leaf curl virus and Yunnan tomato leaf curl virus can also be obviously inhibited.

The invention discloses a rice Os04g0650800 gene promoter and application thereof, belongs to the technical field of plant biology, and successfully constructs a yellow fluorescent protein gene recombinant expression vector pPGDH-YFP driven by the promoter of the gene by cloning the promoter sequence of the Os04g0650800 gene. Agrobacterium tumefaciens infects Nipponbare rice, and positive rice plants are screened. Xanthomonas oryzae pv.oryzae, Xoo is inoculated to the positive rice seedlings of the generation T0, a confocal microscope is used for observation, and it is found that YFP expression can be induced by infection of the positive rice plants with Xoo. Genetic engineering modification is carried out on rice through the promoter, for example, high-efficiency expression of a target gene is regulated and controlled through the promoter, and a rice plant responding to Xoo infection will be obtained; if the promoter is used for driving the expression of disease-resistant related genes in rice, rice plants resistant to bacterial blight can be obtained.

The present invention discloses an application of the PUB62 gene in regulating plant resistance to geminiviruses and a method for cultivating transgenic plants, wherein the geminiviruses are soybean yellow leaf curl virus, tomato yellow leaf curl virus, and Yunnan tomato leaf curl virus. In the present invention, the PUB62 gene of Nicotiana benthamiana is cloned and constructed into the NbPUB62-YFP vector carrying a yellow fluorescent protein. Through the agrobacterium-mediated transformation method, the NbPUB62-YFP vector is introduced into plant cells, and through cell and tissue culture techniques, NbPUB62 overexpressing transgenic plants are obtained. The NbPUB62 overexpressing transgenic plants can significantly inhibit the infection of soybean yellow leaf curl virus, and can also significantly inhibit the infection of tomato yellow leaf curl virus and Yunnan tomato leaf curl virus.

The present invention relates to a chemically modified small interfering RNA (siRNA), and more particularly, to a chemically modified siRNA having enhanced nuclease degradation resistance and effectively exhibiting RNAi activity compared to an unmodified siRNA in which 15 bases are extended at the 5'terminal of a sense strand and the 15 bases are simultaneously subjected to phosphorothioate modification, i.e., a non-bridged oxygen atom of a phosphodiester bond is substituted with a sulfur atom. The invention provides a siRNA delivery tool, a method for promoting siRNA delivery to cause gene silencing and improving the efficiency of delivering siRNA to plant cells and application of the siRNA delivery tool, siRNA modified by thiophosphate can be effectively introduced into YFP (Yellow Fluorescent Protein) transformed arabidopsis thaliana leaves, and gene silencing of YFP is realized. The invention also can effectively silence endogenous genes of tobacco. Therefore, the thiophosphate modified siRNA can be used for silencing a certain gene of a plant to achieve the effect of regulating good growth of the plant or treating related plant diseases and insect pests, and has potential agricultural production value and wide application prospect.

This invention discloses the promoter of the rice Os04g0650800 gene and its applications, belonging to the field of plant biotechnology. By cloning the promoter sequence of the Os04g0650800 gene, a recombinant expression vector pPGDH-YFP driving the yellow fluorescent protein gene was successfully constructed. Positive rice plants were screened by infecting Nipponbare rice with Agrobacterium. The T0 generation of positive rice seedlings were inoculated with Xanthomonas oryzae pv. oryzae. Xoo Using a confocal microscope, it was discovered that... Xoo Infection of positive rice plants can induce YFP expression. Genetic engineering of rice using this promoter, such as regulating the efficient expression of target genes through this promoter, will yield positive results. Xoo Infected rice plants; if this promoter is used to drive the expression of disease resistance-related genes in rice, rice plants resistant to bacterial blight will be obtained.

The invention discloses an optimized promoter PyqjfC55 and an application of the optimized promoter PyqjfC55 in preparation of a high-temperature-resistant microbial sensor. A promoter yqjfC55 is mutated by an error-prone PCR (Polymerase Chain Reaction) method to obtain an optimized promoter PyqjfC55, the optimized promoter PyqjfC55 is connected with yellow fluorescent protein EYFP through fusion enzyme, and finally, the high-temperature-resistant and high-sensitivity microbial sensor capable of sensing explosive molecules 2, 4-DNT in a high-temperature environment is obtained. The optimal culture temperature of the Escherichia coli host is 37 DEG C, the effective working temperature is 42 DEG C, and the Escherichia coli host has the capability of maintaining metabolic activity in a high-temperature detection environment. The biosensor provided by the invention realizes high-sensitivity detection of explosive molecules in a high-temperature environment, improves the sensitivity of the detector for detecting the explosive molecules, and expands the application scene.

The present invention discloses an optimized promoter PyqjfC55 and its application in the preparation of a high-temperature-resistant microbial sensor. In the present invention, the promoter yqjfC55 is mutated by error-prone PCR to obtain the optimized promoter PyqjfC55, and the yellow fluorescent protein EYFP is ligated by a fusion enzyme, and finally a high-temperature-resistant and highly sensitive microbial sensor capable of sensing the explosive molecule 2,4-DNT in a high-temperature environment is obtained. The optimum culture temperature of the Escherichia coli host in the present invention is 37 °C, and the effective working temperature is 42 °C, and it has the ability to maintain metabolic activity in a high-temperature detection environment. The biosensor provided by the present invention realizes the high-sensitivity detection of explosive molecules in a high-temperature environment, improves the sensitivity of the detector for detecting explosive molecules, and expands the application scenarios.

This invention belongs to the field of biosensor technology, specifically relating to a fusion protein, a 2-ketoglutaric acid biosensor, and its applications. Specifically, this invention combines the MifS extracellular sensing domain MifS-SD of the MifS / MifR two-component signal transduction system in Pseudomonas aeruginosa PAO1 with a superfolded circular rearranged yellow fluorescent protein, successfully constructing a 2-ketoglutaric acid biosensor with a high response amplitude. Experiments demonstrate that this biosensor can achieve real-time, in-situ detection of 2-ketoglutaric acid in microorganisms and mammalian cells, and can support the detection of related enzyme activities and the screening of inhibitors in a highly efficient, low-cost, and high-throughput manner, thus possessing good practical application value.

The invention belongs to the technical field of biological detection, and particularly relates to a glutaric acid biosensor based on cyclic rearrangement fluorescent protein and application of the glutaric acid biosensor. According to the invention, a glutaric acid specific transcriptional regulation factor CsiR in pseudomonas putida KT2440 is combined with cyclic rearranged yellow fluorescent protein, so that a glutaric acid biosensor with high performance is developed; it is proved that the biosensor is suitable for accurate, efficient and high-throughput detection of glutaric acid in different biological samples such as bacteria, cell culture media, human serum and urine, and real-time and in-situ detection of glutaric acid in mammalian cells and in different subcellular areas can be achieved. The glutaric acid biosensor has the advantages of high response amplitude, strong fluorescence brightness, proper affinity and the like, and has a wide application prospect.

The invention relates to the technical field of biological detection, and particularly discloses a fusion protein and application thereof in preparation of a phosphatidic acid biosensor. The fusion protein comprises circulating arrangement fluorescent protein, Nir1-N protein located at the N end of the circulating arrangement fluorescent protein and Nir1-C protein located at the C end of the circulating arrangement fluorescent protein. The amino acid sequence of the Nir1-N protein is as shown in SEQ ID NO. 1, and the amino acid sequence of the Nir1-C protein is as shown in SEQ ID NO. 2; the cyclically arranged fluorescent protein is selected from cyclically arranged green fluorescent protein, cyclically arranged yellow fluorescent protein, cyclically arranged Venus fluorescent protein and cyclically arranged blue fluorescent protein. The Nir1-N and the Nir1-C in the fusion protein can be combined with phosphatidic acid, and when the phosphatidic acid is combined with the Nir1-N and the Nir1-C, the fluorescent protein is circularly arranged to generate conformational change, so that a fluorescence signal is changed, and detection is realized.

The invention belongs to the technical field of gene engineering, and particularly relates to an indole-3-acetic acid biosensor as well as a construction method and application thereof. The invention relates to an indole-3-acetic acid sensing plasmid. The plasmid carries a gene for coding a repressor protein, a promoter fragment specifically controlled by a transcription factor, a yellow fluorescent protein gene and a biosensor plasmid vector skeleton. The core of the invention lies in that the sensor has the advantages of high sensitivity, wide dynamic range and low background leakage, and can specifically respond to indole-3-acetic acid and various indole derivatives. The main application of the sensor is that the sensor is integrated into escherichia coli engineering bacteria for producing indole-3-acetic acid, so that high-yield strains can be quickly and efficiently screened from massive mutant libraries, and a key tool is provided for solving the industrialization bottleneck of indole-3-acetic acid microbiological method production.

The invention discloses a high-throughput screening method of a 2-keto-3-deoxy-D-xylonic acid dehydratase mutant library and an application of a mutant in biological synthesis of alpha-ketoglutaric acid. The directed evolution modification method comprises the following steps: carrying out fusion expression on a 2-ketone-3-deoxy-D-xylonic acid dehydratase coding gene (xylX), a Rex protein coding gene (rex) and an annularly arranged yellow fluorescent protein coding gene (cpYFP); the method comprises the following steps: firstly, carrying out primary screening on a 2-ketone-3-deoxy-D-xylonic acid dehydratase random mutant library by utilizing a flow cytometry technology, and screening according to a cell fluorescence value to obtain a mutant which possibly has relatively high catalytic activity; then, re-screening the mutants obtained by primary screening through a deep-well plate ultraviolet spectrum method, so as to obtain mutants with obviously improved catalytic activity; and finally, carrying out shake-flask culture on the mutant obtained by secondary screening, and determining the concentration of extracellular alpha-ketoglutaric acid by adopting a high performance liquid chromatography so as to determine the activity of the 2-keto-3-deoxy-D-xylonic acid dehydratase mutant. The high-throughput screening method disclosed by the invention is high in universality, high in sensitivity, simple, effective and high in throughput, and the screened 2-keto-3-deoxy-D-xylonic acid dehydratase mutant can be applied to biosynthesis of alpha-ketoglutaric acid.