345results about How to "Proliferate fast" patented technology

This invention relates to pork liver cell immortalities and its producing method. Procedures are showed: high live ratio fresh original pork liver cell is got by dispase-collagenase perfusion method; it is cultured for 24 hours. Then cell upper heat removing of recombination retronituse that contains SV40 big T antigen is used to infect the pork cell under condition of polybrene which concentration is 8ug / ml. Then it is medicine pressure filtrated by 500ug / ml G418 after one week, single clone cell is picked for large culture when the cell clone appears and grows to 1.0-2.0cm to get pork cell that can passage. Reinfection is done under condition of 8ug / ml polybrene. Then it is medicine pressure filtrated by 2ug / ml puromycin after one week. Single clone cell is picked for large culture when the cell clone appears and grows to 1.0-2.0cm to get immortality pork cell.

The invention relates to the field of microbes, and particularly discloses a Lactobacillus paracasei strain and application thereof. The collection number of the Lactobacillus paracasei strain is CGMCC No.8903; and the Lactobacillus paracasei strain is separated from feces of children, and belongs to Lactobacillus paracasei subspecies. The strain has the advantages of high proliferation speed and excellent acid production capacity, has the capacities for secreting extracellular polysaccharides and resisting bile salt and gastric acid, is capable of adjusting immunoreaction and inhibiting allergy and inflammation, and is very suitable for preparing food, health-care products, medicines, food replenishers and other products.

The invention provides to a method for tissue culturing and quick propagation of sugarcane with an intermittent immersion bioreactor, which is characterized in that: the method performs propagation culture and rooting culture by transplanting detoxicated sugarcane plants, serving as materials, cultured by generations from induced sprouts of sugarcane stem tip tissues into the intermittent immersion bioreactor. In the intermittent immersion bioreactor, a culture temperature is 29 DEG C; the illumination intensity is 1,500 Lx and the illumination time is 16 hours; an intermittent culture condition is to culture for 3min every 3h and to root for 3min every 6h; the first generation can be propagated to 40 times; after rooting is completed, a proper hardening plantlet treatment is performed; and plantlets are transplanted and the survival rate is about 85 percent. The set of system is high in automation degree, has a much higher multiplication rate than that of a traditional tissue culture method, saves labour cost and increases economic benefits.

The invention discloses a preparation method of a CAR-T cell for treating AIDS-related lymphoma. A CD8+ T cell is used to produce the CAR-T cell, and the CD8-T cell derives from a cord blood T cell. The method includes the following steps: preparing cord blood mononuclear cells from cord blood, removing tumors and other cells by using a human T cell purification kit to obtain T cells, and carryingout separation by using magnetic beads to obtain the CD8+ T cell; activating the CD8+ T cell by using an appropriate medium and appropriate stimulation conditions; transferring CAR to the CD8+ T cellby using lentivirus to prepare the CAR-T cell; and amplifying the CAR-T cell in vitro by using cytokines and activating stimulators to achieve the desired effective dose. The invention also relates to the CAR-T cell prepared by the method, and an application thereof. The CAR-T cell prepared in the invention has a good cell activity and a high proliferation speed, and reduces the attack risk of GVHD; and only the CD8+T cell of the umbilical blood T cells of AIDS patients is used to prepare the CAR-T cell for, so the risk of input CAR-T infected with in-vivo HIV is reduced.

The invention relates to the field of wastewater treatment, and particularly discloses a functional microbial agent for treating livestock and poultry breeding wastewater and a method for preparing the functional microbial agent. According to the scheme, the functional microbial agent and the method have the advantages that ammonia and nitrogen in the livestock and poultry breeding wastewater can be effectively decomposed and transformed by aerobic denitrifying bacteria in the functional microbial agent, and accordingly the contents of the ammonia and the nitrogen in the livestock and poultry breeding wastewater can be reduced to a great extent; synergistic effects can be realized by saccharomycetes, bacillus, lactic acid bacteria and photosynthetic bacteria, COD (chemical oxygen demand) in the livestock and poultry breeding wastewater can be decomposed by different levels, and a large quantity of COD in the livestock and poultry breeding wastewater can be effectively and quickly degraded; the COD, the ammonia and the nitrogen in the livestock and poultry breeding wastewater can be effectively reduced by the functional microbial agent.

The invention relates to the technical field of agriculture and discloses a microbial type mineral fertilizer and a preparation method of the microbial type mineral fertilizer. The microbial type mineral fertilizer disclosed by the invention comprises the following components by weight: 35-70% of medical stone powder, 10-25% of microbial strains and 20-30% of humic acid or humate or mixture of the humic acid and the humate. According to the invention, a novel microbial type mineral fertilizer is prepared by taking microbial strains (such as bacillus subtilis, bacillus jelly and the like), a mineral fertilizer and humic acid (humate) as raw materials, and the fertilizer can accelerate the multiplication rate of the microbial strains, realize slow release and controlled release of secondary and trace elements and regulate soil nutrients, thereby reducing the incidence rate of diseases of crops and improving the yield of crops.

The invention provides a high-protein microbiological feed and a preparation method of the high-protein microbiological feed. Dross is one of products produced in the process of floating and clarifying the mixed juice during sugar preparing by the sugarcane, and is rich in various organic non-sugar components; the dross is always considered as impurities and mixed in the mud juice by a sugar plant, and finally becomes lime sludge to cause waste. The preparation method provided by the invention comprises the following steps of taking the dross in the crane sugar plant as the raw material, and taking a non-protein nitrogen source, phosphate and magnesium salt and the like as the auxiliary materials; mixing the raw material and the auxiliary materials; sterilizing; adding beneficial zymocyte preparation; and fermenting at the aerobic condition so as to obtain the high-protein microbiological feed. By adopting the preparation method provided by the invention, the problems that a large quantity of dross is in the crane sugar plant and is adhered and hardly processed, and the loss of the sugar is serious can be solved well; and the current situation that the protein feed resources are in a shortage can be relieved. According to the preparation method, the cost is low, the period is short, the production technology is simple, the limitation from the field and equipment is less, and the quality and the yield of the high-protein microbiological feed are high, and the preparation method plays an important role in popularization and application of the mixed juice floating and clarifying technology in sugar plants.

The invention provides a kit for separated culture of DC-CIK cells, and an application thereof. The kit comprises a lymphocyte separation liquid, a peripheral blood sample treatment liquid, a CIK cell induced propagation system, a DC induced propagation system, a cell culture bottle coating system, a lymphocyte culture medium GT-T551 and a cell culture bag. The application of the kit in separating and culturing the DC-CIK cells comprises the following steps of separation of peripheral blood sample mononuclear cells, separation of the DC cells and CIK cells, induced propagation of the DC cells, the induced propagation of the CIK cells and co-culture of the DC cells and CIK cells. The lymphocytes obtained from the separation by the kit have very high purity and activity; the propagation rate of the CIK cells is fast; the proportion of CD3+CD56+ double positive cells is high; operation method is simple; and conditions are easy to control. The kit can be widely applied in the separated culture for the DC-CIK cells of human and mammals.

The invention aims to provide an efficient culture method of CIK cells. The prepared CIK cell has the characteristics of high multiplication speed, large cell quantity, high cell viability, high killing capacity on cancer cells and the like. The efficient culture method mainly characterized in that a culture flask coated with laminin and fibronectin is used for culturing PBMCs (peripheral blood mononuclear cells), and the cells can be in a half-adherence state by means of the absorption effect of the two kinds of protein, accordingly, the cells can be stimulated by various factors, and the two kinds of protein can promote CIK cell multiplication; and with the adoption of an anti-CD28 monoclonal antibody, the co-stimulation signal of a T cell subset in the CIK cells can be stimulated, and the activation of the CIK cells is promoted. So far, a research report on jointly using laminin, fibronectin and the anti-CD28 monoclonal antibody to prepare the CIK cells is absent. The method for increasing the cell number and improving the killing activity by adding the three factors during preparation of the CIK cells is firstly provided.

The invention provides a canine parvovirus strain CPV-YH and applications thereof, and belongs to the technical field of microbes. The preservation number of the provided canine parvovirus strain CPV-YH is CGMCC No.12990. The invention also provides applications of the canine parvovirus strain CPV-YH in the preparation of canine parvovirus vaccines and canine parvovirus diagnostic reagents. Based on the canine parvovirus strain CPV-YH, the invention also provides a vaccine composition for preventing and / or treating diseases caused by canine parvovirus and a reagent for detecting a canine parvovirus strain and / or diagnosing diseases caused by canine parvovirus. A novel content and direction are provided for the research on the variation of epidemic virus; a novel strain is provided for the canine parvovirus vaccine development; and a high quality raw material and choice are provided for the vaccine development.

An asexual tissue culture method for reproduction of China fir includes such steps as induced culturing to obtain the bud of fir, inoculating it in the reproductive culture medium prepared from MS (1-1 / 2), 6-BA (0.5-1.0 mg / L) and NAA (0.2 mg / L), and culturing at 23+ / -2 deg.C under illuminance of 1600-2200 Lx. Its advantages are short period (35-45 days), high reproductivity and high survival rate.

The invention discloses a rapid propagation method for suspension cell culture of Millettia speciosa Champ. According to the rapid propagation method, young and tender cotyledons taken from Millettia speciosa Champ pods 50-60 days after flowering are utilized as culture materials, subcultured callus tissue is obtained through inducing culture and subculture, and a stable Millettia speciosa Champ suspension cell culture fluid is obtained through the suspension cell culture. With the adoption of the method, stable Millettia speciosa Champ suspension cells with good quality and high propagation yield can be obtained, and a novel way and method are provided for development and utilization of effective components of the Millettia speciosa Champ and creation of new germplasm resources from the Millettia speciosa Champ with biotechnology methods such as somatic hybridization, genetic transformation, mutant induction and the like.

The invention relates to a method for processing set-style yogurt, which selects inulin in certain proportion to be added into a raw material milk to obtain the good-quality set style yogurt with excellent aroma, flavor and color through material preparing, homogenizing, sterilizing, cooling, leaven inoculating, canning, leavening, cooling, post maturing and other process steps. Because of addition of the inulin, stability of the yogurt colloid structure can be improved, and the quality of the set-style yogurt can be improved.

The invention discloses a preparation method and an application of a controllable CD20 chimeric antigen receptor modified T cell. A Leader-CD20-CD8-4-1BB-CD3 zeta-T2A-HSV-TK gene segment is synthesized and inserted into a pLent-C-GFP vector, lentivirus carrying recombinant pLent-C-GFP-CD20 is packaged, a monocyte induced heterogeneous T lymphocyte is infected with the recombinant pLent-C-GFP-CD20 lentivirus, and the controllable CD20 chimeric antigen receptor modified T cell is obtained. The prepared controllable CD20 chimeric antigen receptor modified T cell has the characteristics of high proliferation speed, tumor recognition mechanism, wide tumor killing spectrum and the like.

The invention discloses a clinical-grade serum-free medium for adherent culture of human neural stem cells. The medium disclosed by the invention comprises a basic medium, basic nutrition additives, plant-based human serum albumins, saccharides, lipid, hormones, antioxidants and related substances for promoting metabolism; the basic medium is prepared from commercial DMEM / F12 and a commercial neurobasal medium according to a ratio of 1:1; the basic nutrition additives comprise insulin, holo-transferrin, apo-transferrin, putrescine, progesterone and sodium selenite; the hormones comprise biotin, corticosterone, lipoic acid, Ve and Ve acetic ester; the antioxidants comprise human-derived catalase, human-derived superoxide dismutase, glutathione and Vc; the related substances for promoting metabolism comprise carnitine, T3 and ethanol amine. The medium can improve a cell expansion speed by two to three times, well keeps stem properties of the neural stem cells, keeps the cell differentiation potential of the neural stem cells and eliminates potential animal-origin endotoxin and viral pollution.

The present invention discloses a feeder-layer-free human pluripotency stem cell culture medium, which contains L-ascorbate-2 magnesium phosphate, sodium selenite, recombinant human insulin, human apotransferrin, basic fibroblast growth factor, transforming growth factor beta 1, heparin lithium and / or heparin sodium, a DMEM / F-12 base culture medium and an osmotic pressure regulator. According to the present invention, with the culture medium, when the hiPSC culture is performed in low density and normal density, the proliferation rate is high, and the cell morphology and the pluripotency are well maintained; the use of the expensive heparitin sulfate is not required so as to substantially reduce the cost; and almost all of the hiPSCs maintaining cultures at the current stage can be met, and various small molecule compounds are added to perform induction reprogramming culture (removal of TGFB1 and DM), such the culture medium is suitable for extensive researches of various basic scientific researches and clinical scientific research experiments.

The present invention discloses the tissue cultivation process for quickly breeding Dysosma versipellis. The process includes selection and processing of explant, inducing culture, proliferating culture, rooting culture, transplanting and other steps. The present invention uses MS culture medium with added auxiliary agents including active carbon, 6-benzyl adenine, gibberellin and corn extract for two key stages including the inducing growth of fascicular seedling and rooting of robust seedling. The present invention solves the technological problems in fast breeding and stable rooting of Dysosma versipellis, and has the advantages of high breeding coefficient, stable technology, short seedling growth period and low cost.

The invention belongs to the method for cultivating economic forests and particularly relates to a method for culture and plant regeneration of an ormosia microphylla in-vitro embryo. According to the method disclosed by the invention, the culture and plant regeneration of the ormosia microphylla in-vitro embryo is completed by adopting embryo induced culture, primary bud inducted culture, bud multiplication culture, strong seedling culture, and root induction culture. The method for culture and plant regeneration of the ormosia microphylla in-vitro embryo is higher in expanding propagation coefficient, short in germination time and high in propagation speed, and has an important theoretical significance and higher economic value as well as development utilization prospect for efficiently nursing rare and endangered ormosia microphylla resources, through the strong seedling culture, a grown-up seedling is high in survival rate after being transplanted, and the seedling is healthy and strong as well as tall and straight with good growth vigor.

The invention relates to asparagus parent breeding method, mainly solving the problems that rooting rate is low, root quality is poor and the survive rate of the regeneration plant is low in asparagus organ tissue culture process. The method includes the process that high quality asparagus stem section is selected, culture is carried out in culture medium solution, and asparagus seedling meeting transplant standard is bred. The culture medium solution is confected by the way that hormones corresponding to various culture stages are added on the basis of the basic culture medium solution, the culture includes induction culture, propagation culture, rooting pre-differentiation culture and rooting culture sequentially. Compared with the existing organ culture, the invention has high propagation rate, rooting rate reaches 100%, root system quality is good, and differentiation rate of strong fleshy tap root is improved to 96%, thus ensuring survival rate after transplantation.

Belonging to the technical field of cell mediums, the invention in particular relates to a serum-free medium for rapid generation of induced pluripotent stem cells. The serum-free medium for rapid generation of induced pluripotent stem cells includes a basic medium and an additive, and the additive comprises the following components in terms of final concentration: 20-40ng / mL cell growth factor, 21-30microg / mL transferrin, 6-9microg / mL fibronectin, 10-15microg / mL polyvinylpyrrolidone, 1-3microg / mL genistein, 4-7microg / mL mannitol, 5-8microg / mL insulin, 11-15microg / mL L-glutamine, 12-16microg / mL palmitic acid, and 8-15microg / mL hesperidin. The serum-free medium for rapid generation of induced pluripotent stem cells provided by the invention has the advantages of clear components, no serum, high safety, and rapid generation of induced pluripotent stem cells.

The invention provides an immortalized dairy cow rumen endothelial cell line and a construction method thereof. The construction method comprises the following steps: (a) cutting, washing, and digesting collected dairy cow rumen endothelial tissues in a culture medium, and carrying out primary culture to obtain rumen endothelial cells; step (b) incubating rumen endothelial cells obtained in the step (a) with a viral liquid containing an SV40T antigen gene to obtain infected rumen endothelial cells; and step (c) culturing the infected rumen endothelial cells obtained in the step (b) to obtain the immortalized dairy cow rumen endothelial cell line. The provided immortalized dairy cow rumen endothelial cell line can provide a test cell model for the research on physiological regulation and nutrient absorption mechanism of dairy cow rumen. The culture method is simple, the growth speed is quick, and the construction method can obtain BRECs, which have physiological functions and can carryout continuous passage.

The invention discloses a method for synchronously removing nitrate and arsenic in underground water by natural pyrrhotite and an application of the method, and belongs to the technical field of water resource purification. The method includes the steps: (a) material preparation: crushing the natural pyrrhotite into particles and washing the particles into neutral with water after acid pickling; (b) strain screening: screening sulfur-based autotrophic denitrification bacteria taking the natural pyrrhotite as a sulfur source from anaerobic sludge by a specific screening medium to serve as a target strain; (c) sewage treatment: taking the underground water containing the nitrate and the arsenic, inoculating the target strain and then placing the underground water in a reactor containing the natural pyrrhotite for reaction; (d) solid-liquid separation. The natural pyrrhotite is used as the sulfur source of the sulfur-based autotrophic denitrification bacteria for denitrification, the nitrate in the water is transformed into nitrogen in a reductive manner, the pyrrhotite and oxidation product thereof can remove the arsenic in the water by the aid of functions such as chemical precipitation and adsorption, so that the nitrate and the arsenic in the underground water are synchronously and efficiently removed. The method is convenient in operation and low in cost.

The invention discloses a special foliar microbial bacterium fertilizer for paddy rice, and relates to the technical field of microorganism. The microbial fertilizer comprises, by mass, 4-6 parts of potassium fulvate, 2-4 parts of a bacillus agent, 0.1-0.25 part of potassium dihydrogen phosphate, 0.2-0.8 part of potassium sulfate, 0.03-0.08 part of magnesium sulfate, 0.1-0.25 part of diammonium phosphate, and the balance of water; and microbial bacteria of the bacillus agent include Bacillus subtilis and Bacillus methylotrophicus. The invention also provides a preparation method of the specialfoliar microbial bacterium fertilizer for paddy rice. The preparation method of the microbial fertilizer comprises the following steps: preparation of the bacillus agent, and preparation of the microbial fertilizer. The microbial fertilizer provided by the invention is applied to the control process of rice blast, and the strain can be stably colonized in various parts of crops and occupy the ecological niche of the pathogenic bacteria, so the growth of the pathogenic bacteria is inhibited, the growth of the crops is promoted, the yield of the crops is increased, and the product quality is improved.

The invention discloses a quick propagation method for humulus scandens. The method comprises the following steps of: culturing tissues, such as stem section of the humulus scandens, on an LS culture medium; accurately cutting and taking the stem section with 1-2 axillary buds as an explant; disinfecting and then inoculating in an LS+6-BA 1.2mg / L+NAA 0.2mg / L culture medium for culturing; wherein the explant buds after being inoculated for about 6-8 days and grows into a plantlet with leaves after being inoculated for about 10 days; cutting off the plantlet and transferring to an LS+6-BA 0.5mg / L+IAA 0.4mg / L culture medium, so as to keep high-speed increasing and a propagation coefficient above 5; selecting LS+6-BA 0.5mg / L+IAA 0.4mg / L+NAA 0.5mg / L as a rooting culture medium, wherein the rooting rate is above 90% and the complete plant can be obtained only after about 25 days; and transplanting a test-tube plantlet into sandy soil, carefully keeping water and humidity, and establishing a set of high-frequency stable regeneration system after the transplanting survival rate is above 90% after transplanting for 1 month. The method provided by the invention has the advantages of excellent stability, convenience in operation, high propagation speed and low production cost, achieves industrial level, and the like.

The invention discloses a recombinant serine protease inhibitor, a fungus expression vector and a fungus insecticide of recombinant protein. The invention relates to the recombinant serine protease inhibitor; a molecular biology technology is used to construct a recombinant nucleotide sequence; and the recombinant serine protease inhibitor comprises a signal peptide sequence of encoded coccidio beauveriabassiana chitinase and a serine protease inhibitor gene of encoded drosophila, and has a nucleotide sequence shown as SEQ ID NO. 1 and an amino acid sequence shown as SEQ ID NO. 2. The obtained recombinant serine protease inhibitor can inhibit immunoreaction of insects, increase multiplication speed of fungi in haemocoeles, and therefore improve characteristics of insecticide fungus toxicity.

The invention relates to a novel microorganism flora mixture and a mixed nutrient medium thereof. The nutrient medium is filter liquor which is prepared by taking plant powder, mineral composition, animal powder, organic acid and the like as raw materials, adding sewage or comprehensive sewage in cities to be processed, mixing, fermenting and filtering. The microorganism flora mixture is a microorganism preparation which is prepared by the following steps: respectively selecting fusarium oxysporum, aspergillus fumigatus, sphaerotilus, vorticella microstoma, photosynthetic bacteria, methanebacteria, desulphovibrio, nitrobacterium, pseudomonas fluorescens and other bacterial strains from obligate aerobes group, anaerobic bacteria group and facultative aerobe group preferentially, then expanding and cultivating, inoculating into the above mixed nutrient medium, mixing, cultivating and domesticating for 15-30 days at the temperature of 20-45 DEG C. The mixed nutrient medium in the invention can cultivate, propagate and domesticate flora mixture with the combination advantages of flora; the flora has stable integrity, no easy variation and strong adaptability, and the invention breaks through the technical bottlenecks of easy degeneracy of microorganism flora, easy decrease of biological activity and slow proliferation and expansion.

The invention discloses a tissue culture rapid propagation method of callicarpa nudiflora. The method performs control on the explant collection, disinfection method, differentiation, multiplication, screening of a rooting medium, selection of a transplanting matrix, environmental conditions in the culture process and the like. By adopting the methods of explant selection, inoculated culture, propagation expanding culture, rooting culture and hardening-seedling transplanting, a set of complete callicarpa nudiflora tissue culture rapid propagation technology system is established, so that the differentiation rate reaches 100%, the monthly multiplication coefficient reaches 4.95, the rooting rate reaches 93.7%, and the transplanting survival rate exceeds 90%. The method disclosed by the invention is simple and easy to implement, simple in process and high in efficiency, effectively suppresses the pollution and browning of the callicarpa nudiflora explant, improves the differentiation rate of the tissue culture seedling, realizes fast multiplication, high rooting rate, high survival rate and low cost, and provides a reliable technical means for large-scale production of callicarpa nudiflora seedlings.

The invention belongs to the field of culture mediums, and in particular relates to a T cell serum-free medium and a using method thereof. The T cell serum-free medium consists of a basal culture medium applied to growth culture of cells, as well as the following addition ingredients: ethanolamine, copper sulfate, ferric nitrate, zinc sulfate, sodium selenite, sodium pyruvate, insulin, transferrin, glutamine, serum albumin, thioglycerol and L-vitamin C. The T cell serum-free medium provided by the invention breaks through shortcomings of the prior art, and according to the T cell serum-free medium, the entire medium system is free from the introduction of non-human proteins, and the T cells in the blood sample, which is safe and reliable in source, undergo efficient selective amplification; and therefore, higher clinical application and scientific research values are achieved.

The invention provides a method for preparing an ionic rare earth mine tailing in-situ remediation plant nursing system. The method utilizes agricultural waste as a biomass carrier and a Bacillus amyloliquefaciens H47 strain as a plant growth promoting inocula which is preserved in the China center for type culture collection on June 24, 2013 and has a preservation number of CCTCC NO: M2013283 to prepare the ionic rare earth mine tailing in-situ remediation plant nursing system. Through immobilization of the biomass carrier, activity of the plant growth promoting inocula can be kept for a long time and the plant growth promoting inocula can produce stable and efficient action in a complex environment. The system can improve soil fertility and prevents soil hardening and fast soil moisture evaporation. The system utilizes the agricultural waste biomass carrier and has a wide source, a low cost and simple processes. The system realizes waste utilization, can be used as bio-fertilizer, and provides various nutrients for a plant. Through the system, the restored soil is suitable for plant growth.

The invention provides a novel composite probiotic preparation for a pig-raising fermentation bed, wherein the preparation comprises the following components: bacillus subtilis, anaerobic bacteroid, lactobacillus fermenti, bifidobacteria, lactobacillus, saccharomycetes, fructooligosaccharide, isomalto-oligosaccharide, DL-methionine, L-lysine, multi-vitamin, and soybean powder. The probiotics, which are used for the preparation of the ecological pig-raising fermentation bed, propagate and grow well in organic materials such as straw, rich husk, and the like, can strengthen the decomposition efficiency of pig manure and urine, have a stable number of bacterial colonies in the culture period, effectively prolong the service life of the fermentation bed, at the same time help to modulate the microecological balance in the pig intestinal tracts, improve the immunity function and productive performance of ecologically-fed pigs, and prominently increase the economic profit of pig feeding.