128 results about "Nicotiana benthamiana" patented technology

Disclosed herein are GNGN and G1 / G2 antibodies that recognize and bind various FcRs and C1q. Also disclosed herein are glycan-optiminzed antibodies, predominantly of the GNGN or G1 / G2 glycoform, with enhanced Fcγ receptor binding achieved through CHO, Nicotiana benthamiana and yeast manufacturing systems. Nucleic acids encoding these antibodies, as well as expression vectors and host cells including these nucleic acids are also disclosed herein. Methods and pharmaceutical compositions including the monoclonal antibodies are provided herein for the prevention and / or therapeutic treatment of viral infections, cancers and inflammatory diseases.

The invention relates to an application of plant eIF4A genes in preparation of rice stripe virus infection resistant rice. According to the invention, a binary expression vector is constructed by utilizing the plant eIF4A genes and vsiRNA binding site mutants thereof; rice and nicotiana benthamiana are successfully converted; and a rice stripe virus infection resistant transgenic plant is obtained. Experimental results show that the tolerance of the plant body to rice stripe virus can be obtained through expression of the plant eIF4A genes or the vsiRNA binding site mutants thereof in the plant body.

The invention discloses a tobacco PDS gene fragment-containing cucumber mosaic virus RNA2 attenuated mutant type plasmid vector and application thereof. The plasmid vector contains a CMVFny strain RNA2 complete sequence, and a tobacco PDS gene 199bp fragment is inserted behind a 2a protein termination codon. The plasmid vector and a wild CMVFny RNA1 and wild CMVFny RNA3-containing plasmid are inoculated to nicotiana benthamiana through an agrobacterium infiltration method, so that weakly pathogenic symptoms can be displayed, and the capacity of inducing gene silencing can be judged with nakedeyes without depending on an instrument; great convenience is provided for deep researches on attenuated vaccines.

The invention relates to a grapevine powdery mildew resistance transcription factor gene VpRFP1 promoter sequence and application thereof. In the sequence, a full-length 1249bp promoter sequence of a Chinese wild vitis pseudoreticulata powdery mildew transcription factor gene VpRFP1 is cloned by adopting chromosomal walking in combination with a nested polymerase chain reaction (PCR) technology, wherein the sequence comprises 103 TATA-boxes and 27 CAAT-boxes, has 8 elements related to plant defense reaction, 14 photoreaction elements, 5 plant hormone response elements, and 14 other unknown elements. By adopting an agrobacterium tumefaciens-mediated instantaneously converted nicotiana benthamiana leaves analyzing method, the function of the Chinese wild vitis pseudoreticulata powdery mildew transcription factor gene VpRFP1 promoter proves that: the promoter has stronger promotion activity, can actively respond to pathogen invasion and disease resistant signal substances, and can improve the disease resistance and high temperature resistance of wheat, rice, potato, corn, soybean, rape and other plants through a transgenic method.

Plant viral vectors have great potential in rapid production of proteins, but no simple. Here a geminivirus-based system for high-yield and rapid production of oligomeric protein complexes, including virus-like particle (VLP) vaccines and monoclonal antibodies (mAbs) is described. In particular, a single vector that contains two non-competing replicons for transient expression in Nicotiana benthamiana leaves is described. The correct assembly of these subunit proteins into functional oligomeric structures (VLPs or full-size mAb) is also described. This system advances plant transient expression technology by eliminating the need for non-competing viruses, and thus, enhances the realistic commercial application of this technology for producing multiple-subunit protein complexes.

The invention discloses an obtaining method and application of a soybean mosaic virus infecting tobacco, and an application of the soybean mosaic virus infecting tobacco. A soybean mosaic virus isolate having a preservation number of CCTCC NO:V201349 is used to invade Nicotina benthamiana to obtain the soybean mosaic virus infecting tobacco. The soybean mosaic virus infecting tobacco obtained through the method can be applied in the verification of anti-soybean mosaic virus candidate genes. The soybean mosaic virus capable of invading tobacco is obtained by screening from many isolates, and confirmation is carried out through the isolate invading tobacco, disease symptom investigation, Nannong 1138-2 backtie experiment, ELISA and an RT-PCR detection means. The soybean mosaic virus infecting tobacco is obtained in the invention, realizes the verification of the anti-SMV candidate gene function on the tobacco, and breaks the restriction that the genetic transformation in soybean is difficult.

A single vector or multiple separate vectors that contain two or more non-competing replicons for transient expression of the heavy and light chains of Rituximab in Nicotiana benthamiana leaves is described. The correct assembly of these subunit proteins into functional oligomeric structures to optimize the expression is also described. This system advances plant transient expression technology by eliminating the need for non-competing viruses, and thus, enhances the realistic commercial application of the multi-replicon single vector system for producing Rituximab in plant cells.

Plant viral vectors have great potential in rapid production of proteins, but no simple. Here a geminivirus-based system for high-yield and rapid production of oligomeric protein complexes, including virus-like particle (VLP) vaccines and monoclonal antibodies (mAbs) is described. In particular, a single vector that contains two non-competing replicons for transient expression in Nicotiana benthamiana leaves is described. The correct assembly of these subunit proteins into functional oligomeric structures (VLPs or full-size mAb) is also described. This system advances plant transient expression technology by eliminating the need for non-competing viruses, and thus, enhances the realistic commercial application of this technology for producing multiple-subunit protein complexes.

The invention relates to clone, reconstruction and transformation of a precursor (pre-miR159a) of arabidopsis thaliana microRNA159a and antivirus analysis of a transgenic plant and belongs to the technical field of molecular biology and biology. The technology clones a nucleotide sequence of pre-miR159a from the arabidopsis thaliana and carries out site-directed mutagenesis on the sequence producing mature miR159a. 126kDa protein of tobacco mosaic virus, 25kDa protein of potato virus X and HC-Pro protein of potato virus Y are subjected to genetic mutation to obtain three sections of the mutation sequences. The three sections of the sequences are in series to construct the plant expression vector; and an agrobacterium-mediated method is adopted to transform nicotiana benthamiana to obtain transgenic tobacco. The transgenic plant has high disease resistance on three viruses. The antivirus strategy has the advantages of strong disease resistance, durable disease resistance, biological safety and the like and has wide application prospect in the breeding field of plant antivirus genetic engineering.

The invention relates to a gene GhMKK5 capable of improving bacterial resistance of transgenic crops and an application thereof, belonging to the fields of molecular biology and biotechnology. According to the invention, a cotton mitogen-activated protein kinase gene GhMKK5 is obtained by the homologous cloning and RACE technology; the cDNA (complementary deoxyribonucleic acid) of the gene is placed under a CaMV 35S starter to construct a plant expression vector; and nicotiana benthamiana is transformed so that the gene is over-expressed in tobacco. Experiments demonstrate that: the transgenic tobacco has obvious capability in resisting bacteriosis. By applying the gene to the crops such as transformed wheat, cotton and the like, the antibacterial capability can be enhanced, the yield and quality can be improved, and tremendous economic and social values can be realized.

The invention mainly relates to construction of a gene silencing vector induced by Chinese wheat yellow mosaic virus, and application thereof to nicotiana benthamiana. The biological active analysis results show that the constructed CWMV-VIGS can successfully infect the nicotiana benthamiana and can realize the silencing after the effective transcription on relevant genes such as phytoene desaturase and 50S ribosomal protein L12. The types of the VIGS vectors are enriched; in addition, the effective technical measures are provided for the gene function study.

This invention encompasses the identification and isolation genes and gene fragments that confer altered metabolic characteristics in Nicotiana benthamiana plants, when expressed using GENEWARE™ viral vectors. These genes are derived from a variety of sources. Expression of these genes resulted in alterations of the levels of at least one of the following metabolites: acids, fatty acids, amino acids and related compounds, branched fatty acids, carbohydrates, hydrocarbons, alkaloids and other bases, esters, glycerides, phenols and related compounds, alcohols, alkenes and alkynes, sterols, oxygenated terpenes, and other isoprenoids, and ketones and quinones.

The invention relates to a gossypium hirsutum mitogen-activated protein kinas 6 (GhMAPK6) gene and application thereof, belonging to the fields of molecular biology and biotechnology. The GhMAPK6 gene is obtained by adopting homologous cloning and RACE [Rapid-Amplification of cDNA (complementary Deoxyribonucleic Acid) Ends] technology. The cDNA sequence of the gene is put under a CaMV 35S promoter to construct a plant expression vector, and nicotiana benthamiana is transformed to excessively express the gene in tobacco. An experiment proves that a transgenic tobacco plant has remarkably improved yield than a non-transgenic tobacco plant in the aspects of increase in the leaf area, increase in the tillering number, increase in the infruit number and increase in the ripening rate. Therefore, the gene has important realistic significance in improving the crop yield trait and increasing the crop yield by a gene engineering way according to the yield increasing property of the gene.

The invention discloses a ternary shuttle vector pCY which has a nucleotide sequence of SEQ ID NO:17 as shown in the specification. The invention further discloses a method for establishing CLBV (Citrus Leaf Blotch Virus) infectious cloning. The method comprises the following steps: (1) extracting total RNA (Ribonucleic Acid) of an infectious CLBV plant, carrying out reverse transcription, and carrying out PCR (Polymerase Chain Reaction) amplification with pCY-CLBV1F, CLBV1R and CLBV2F and pCY-CLBV2R respectively so as to obtain specific segments CLBV1 and CLBV2 covering the whole length of aCLBV genome; (2) carrying out digestion on pCY with Stu I and Sma I; (3) carrying out TAR (Homologous Recombination) cloning, namely carrying out co-transformation on yeast YPH501 with CLBV1, CLBV2 and linearized pCY by using a lithium acetate method, and carrying out homologous recombination so as to obtain CLBV whole length cDNA (Complementary Deoxyribonucleic Acid) cloned pCY-CLBV; (4) carryingout pCY-CLBV plasmid transformation on agrobacterium, inoculating with citruses or nicotiana benthamiana, and carrying out identification, thereby completing CLBV infectious cloning.

The invention discloses a construction method for simultaneously expressing two foreign protein vectors TRVe2. The construction method comprises the following steps: cloning pYL156 by agrobacterium infection of genome RNA2 of TRV as a material, constructing a vector containing 2b gene promoter and pea early-browning virus coat protein gene promoter sequences of TRV, and driving expressions of foreign genes in nicotiana benthamiana by virtue of genome promoters of the two plant viruses. The plant virus TRV2e2 capable of simultaneously expressing two non-fusion-proteins is first reported internationally; the virus does not cause any obvious symptoms in the nicotiana benthamiana, carries two foreign genes to systematically expand to the whole plant and is capable of expressing two foreign proteins in the same cell.

The invention relates to a method for screening disease-resistant genes by the aid of tomato spotted wilt virus NSm (new smoking material) genes, and belongs to the technical field of plant gene cloning. Nucleotide sequences of the tomato spotted wilt virus NSm genes are shown as SEQ ID No.1. The method includes constructing plant expression vectors of the tomato spotted wilt virus NSm genes; acquiring agrobacterium tumefaciens by means of transformation; jointly infiltrating nicotiana benthamiana in the agrobacterium tumefaciens and candidate disease-resistant genes; observing nicotiana benthamiana leaves to determine whether the nicotiana benthamiana leaves have characters of hypersensitive necrosis or not; authenticating the candidate disease-resistant genes. The method has the advantages that the disease-resistant genes can be quickly authenticated by the method in a high-throughput manner, and accordingly important application value of the method developed on the basis of genes for screening disease-resistant genes of plants is indicated.

Expression vectors and methods of their use for enhancing the production of recombinant proteins in plants or plant cells are described. Production can be further enhanced upon co-expression of the P19 suppressor of gene-silencing from tomato bushy stunt virus. Preferably, the recombinant proteins are therapeutic enzymes and / or antibodies and methods are carried out in Nicotiana benthamiana—optionally an RNAi-based glycomodified strain—or in the Nicotiana tabacum cultivar Little Crittenden.

The invention discloses a potato leaf roll virus infectious clone and a construction method thereof. The potato leaf roll virus infectious clone is obtained by cloning a whole genome of a potato leafroll virus into a pCB301 binary vector containing a cauliflower mosaic virus double 35S promoter in a homologous recombination mode. The potato leaf roll virus infectious clone which can be inoculatedby an agrobacterium infiltration method and stably and efficiently infects host plants such as nicotiana benthamiana and potatoes is prepared for the first time, and the potato leaf roll virus infectious clone has important significance for preventing and treating potato leaf roll virus diseases.

The invention discloses protein BxCDP1 of bursaphelenchus-xylophilus pathogen-associated molecular patterns and application thereof. The amino acid sequence of the protein BxCDP1 of the pathogen-associated molecular patterns (PAMPS) is shown in SEQID NO.2. From effectors secreted by bursaphelenchus xylophilus and the PAMP, the defensive responses of a host plant, namely, a pine tree, to bursaphelenchus xylophilus invasion are studied, the protein BxCDP1 of the pathogen-associated molecular patterns is obtained from the bursaphelenchus xylophilus, it is proven through experiments that the protein BxCDP1 can trigger cell necrosis of various plants including the host plant, has a certain broad spectrum in cell necrosis triggering, and stimulates the defensive response of the host plant. The BxCDP1 triggered cell necrosis depends on a co-receptor BAK1 of pattern recognition receptors, the BxCDP1 can stimulate the accumulation of nicotiana benthamiana ROS and up regulation of PTI Marker genes, and the immunoreaction of the nicotiana benthamiana is stimulated. It can be seen that the BxCDP1 is a PAMP secreted by the bursaphelenchus xylophilus, and has important theoretical and practicalsignificance for revealing the pathogenic mechanism of the bursaphelenchus xylophilus and improving the resistance of the pine tree to the bursaphelenchus xylophilus in a targeted mode.

This invention encompasses the identification and isolation of genes that confer disease control properties in plants, as well as plants comprising such genes. These genes are derived from the following sources: Nicotiana benthamiana, Oryzae sativa (var. Indica IR7), Papaver rhoeas, Saccharomyces cerevisiae and Trichoderma harzianum (Rifai 1295-22). The control conferred is against the one or more of the following phytopathogens: Aspergillus flavus, Cercospora zeae-maydis, Fusarium monilforme, Fusarium graminearum, Helminthosporium maydis, Phoma lingam, Phomopsis helianthi, Phytopthera infestans, Pyricularia oryzae, Pythium ultimum, Rhizoctonia solani, Sclerotinia sclerotiorum, Ustilago maydis, and Verticillium dahliae. Further, this invention encompasses other homologous and heterologous sequences with a high degree of functional similarity.

A single vector or multiple separate vectors that contain two or more non-competing replicons for transient expression of the heavy and light chains of Rituximab in Nicotiana benthamiana leaves is described. The correct assembly of these subunit proteins into functional oligomeric structures to optimize the expression is also described. This system advances plant transient expression technology by eliminating the need for non-competing viruses, and thus, enhances the realistic commercial application of the multi-replicon single vector system for producing Rituximab in plant cells.

The invention relates to a novel cotton fungal disease-resistant gene GhMPK7 and application thereof, which belong to the field of molecular biology and biotechnology. In the invention, a gossypium hirsutum mitogen-activated protein kinases gene is obtained by adopting homology-based cloning and RACE technology. A cDNA sequence of the gene is positioned below a CaMV35S promoter to construct plant expression vectors and convert nicotiana benthamiana so that the gene is excessively expressed in tobacco. Experiments show that transgenic tobacco has high resistance to fungal diseases, in particular to colletotrichum nicotianae and alternaria alternate. When the gene is applied to crops such as wheat, corns and cottons, the fungal resistance can be improved and the yield and the quality can be improved. Thus, the gene has significant economical and social benefits.

The invention discloses a nicotiana benthamiana secreting type glandular hair regulation gene NbJAZ3 as well as an expression vector and application thereof, and aims to solve the technical problems that the generation mechanism of tobacco secreting type glandular hair is unknown and an effective regulation means is lacked at present. The gene NbJAZ3 capable of regulating secreting type glandular hair generation is identified, an overexpression vector and a silence vector of the gene are constructed, and a corresponding overexpression strain and a corresponding gene silence strain are respectively obtained through transformation; and phenotypic and physiological feature identification and detection show that the NbJAZ3 gene can negatively regulate the generation of tobacco secreting type glandular hair, and has important application value in the aspects of glandular hair generation density regulation, glandular hair type directional improvement and the like.

The invention discloses a vector for editing a nicotiana benthamiana gene based on CRISPR / Cas9 as well as a construction method and application thereof. The vector comprises a BKG-g10 vector and a BKG-g11 vector which are constructed by aiming at a nicotiana benthamiana UbEF1B gene, and / or a BKG-g12 vector and a BKG-g13 vector which are constructed by aiming at a nicotiana benthamiana CCR4 / NOT gene. According to the vector, after the UbEF1B gene or the CCR4 / NOT gene is edited, the growth state of nicotiana benthamiana is normal, and the phenotype is not obviously changed compared with that ofa wild type; and no influence is caused on the character, and progeny continuation is facilitated. Compared with the traditional breeding method, the method for editing a nicotiana benthamiana endogenous gene by virtue of a CRISPR / Cas9 system has the advantage that a disease-resistant material can be obtained more quickly and effectively.

The invention belongs to the technical field of gene engineering, particularly relates to a potato U6 RNA polymerase III type promoter, more particularly relates to StU64-1 and StU68-1 gene promoters, and further discloses a cloning method and application thereof. Two potato RNA polymerase III type promoters StU64-1 and StU68-1 are obtained by cloning in potatoes, have efficient transcriptional activity and can drive downstream sgRNA expression, and the activities of the two promoters and the feasibility of applying the promoters to potato CRISPR / Cas9 gene editing are respectively verified by virtue of an instantaneous transformation system of nicotiana benthamiana leaves and a stable transformation system of potatoes, and the CRISPR / Cas9 guided potato genome editing is realized. In the technical field of transgenosis, the promoters are not only suitable for potatoes, but also can be applied to other solanaceae crops such as tobacco.

The invention provides Nicotiana benthamiana mutant plants which are incapable of forming xylosyl-structures on glycoproteins. In addition, the invention provides methods for the production of heterologous glycoproteins in said mutant plants.

The invention discloses an application of a Verticillium dahliae VdPRMT1 gene in improvement of the disease resistance of crops. The method comprises the following steps of: constructing a plurality of tobacco rattle virus interference plasmids aiming at the Verticillium dahliae VdPRMT1 gene by taking a Verticillium dahliae strain as a material through host-induced gene silencing, converting Nicotina benthamiana, inoculating the Verticillium dahliae, and preliminarily determining that the VdPRMT1 gene in the Verticillium dahliae is associated with pathogenicity. The Verticillium dahliae VdPRMT1 is further taken as a target gene, and a target gene segment capable of remarkably improving the plant disease resistance is obtained through screening by combining transgenosis and an RNAi (Ribonucleic Acid Interference) technology. The screened target gene segment and a Gateway interference vector constructed by applying the target gene segment can be applied to the aspects of improving the resistance of plants to diseases caused by the Verticillium dahliae, cultivating new varieties of transgenic plants capable of resisting the Verticillium dahliae and the like.

The invention discloses a verticillium dahliae phospholipase target gene fragment and an interference vector and application thereof, and belongs to the fields of clone and application of verticillium dahliae disease course key genes. A gene sequence of a verticillium dahliae phospholipase target gene is introduced into transient expression ds RNA of nicotiana benthamiana through a virus-induced gene silencing technology, and the target gene which can extremely significantly improve the plant disease resistance is screened; an RNAi technology is further utilized to establish the stably inherited interference vector and transform the nicotiana benthamiana, and transforming results show that transgenic tobacco plants extremely significantly improve the disease resistance of tobaccos to verticillium dahliae, show the highly resistance and even reach the immune level. The verticillium dahliae phospholipase target gene fragment and the RNA interference vector can be applied to improving the disease resistance of the plants to the verticillium dahliae and cultivating new transgenic plant species with the verticillium dahliae resistance.

The invention relates to an RNA-RP RT-PCR method capable of fast detecting melon viruses. Leaves of melon plants infected with viruses are homogenized by adopting a PBST plus a 2% PVP solution or a 0.5% Triton X-405 PBS solution, sample virus RNA is extracted in supernate, and the supernate is used for one-step RT-PCR amplification of target segments of the viruses, so that the melon viruses are detected. According to the RNA-RP RT-PCR method capable of fast detecting the melon viruses, only trace amounts of samples are needed for obtaining adequate amounts of RNA templates, the RNA templatesare used for detecting the six viruses of a Cucumber green mottle mosaic virus (CGMMV), a Cucumber mosaic virus (CMV), a Zucchini yellow mosaic virus (ZYMV), a Watermelon mosaic virus (WMV), a papayaring spot virus (PRSV) and a squash mosaic virus (SqMV) in watermelons, muskmelons, cucumbers, zucchini and a model plant nicotiana benthamiana in melon plants, the method is high in sensitivity and accurate in result, operation is simple and fast, and a novel detection method of the melon viruses is provided.