274 results about "Sepharose" patented technology

The invention relates to cordyceps militaris active polysaccharide prepared by a method comprising the following steps: performing ultrafine grinding on cordyceps militaris sporophores, then extracting with boiling water, and collecting supernatant fluid; and concentrating the supernatant fluid, then adding ethanol for deposition, passing a deposited part through a DEAE-sepharose column, desalting a part of 0-0.5N, and then performing freeze-drying. The invention also relates to an application of a cordyceps militaris extract in preparation of an anti-tumor activity product.

The present present invention relates to a method of simultaneously extracting phycoerythrin and agar from seaweeds such as asparagus, etc in seaweeds biological chemical field. The concrete operation process is that cells of the asparagus frond are crushed. A phycobiliprotein crude extract is obtained from water soluble extracts after ammoniym sulfate precipitation. The crude extract is extracted by Phenyl-Sepharose expansion columns and phycoerythrin products are obtained. Q-Sepharose iron exchange columns are used to purify the phycoerythrin products to obtain purified phycoerythrin products. Then, the left asparagus residues after the phycoerythrin extract are used to extract the agar. The present invention can simultaneously obtain the phycoerythrin and the agar from the asparagus. And the purified phycoerythrin purity is larger than 2.954, and the yield is 0.097mg per gram of fresh algae. The yield of the obtained agar is 2.31percent (the algae is wetly weighted), which is equivalent to the yield of an agar direct extraction method. Therefore, the utilization ratio of the asparagus can be greatly improved by adopting the method.

The invention provides a preparation method of flavobacterium heparinum heparinases I, II and III, which comprises the following steps: inoculating flavobacterium heparinum used as the raw material into a seed culture medium for culture, then inoculating into a fermentation culture medium, centrifuging, collecting the precipitate, carrying out ultrasonication on the precipitate, and centrifuging to obtain a crude enzyme liquid of flavobacterium heparinum heparinases I, II and III; carrying out SP-Sepharose FF on the crude enzyme liquid to obtain an enzyme II and enzymes I and III through chromatographic separation; carrying out SP-Sepharose FF on the enzymes I and III to obtain a heparinase I and a heparinase III through separation; respectively carrying out SP-Sepharose FF on the obtained enzyme I and enzyme III twice to obtain high-purity heparinases I and III through purification; and respectively carrying out HEP-Sepharose 4B, HA-Ultrogel and SP-Sepharose FF on the obtained enzymeII to obtain a high-purity heparinase II.

The invention discloses a preparation method for human immunoglobulin for intravenous injection. The preparation method comprises the following steps: (1) two-step filtering pressing: separating II+III precipitate from human plasma as a starting raw material, removing component III precipitate through low-temperature ethanol filter pressing, collecting supernatant, performing ultrafiltration and dialysis to the supernatant, and regulating the protein concentration; (2) two-step chromatography: performing upper column chromatography purification through a DEAF-Sepharose-FF ion exchange column, and collecting a first-step flow penetration liquid; performing ultrafiltration and dialysis to the collected first-step flow penetration liquid, regulating and performing chromatographic purification by using a Fractogel-EMD-TMA ion exchange column, and collecting a second-step flow penetration liquid; (3) performing ultrafiltration, dialysis and liquid preparation to the collected second-step flow penetration liquid; (4) inactivating virus, disinfecting, filtering and subpackaging to obtain a product. The product prepared by using the method is higher in bioactivity, yield and purity.

The invention relates to a method for observing microscopic structures inside plant roots, which is capable of effectively solving the problem of utilizing a microscope to clearly observe the structures inside the plant roots. The method comprises the steps of digging out the plant roots, washing, shearing the plant roots into small sections, fixing the plant roots by methanol, and then embedding by a sepharose solution, cooling, condensing, trimming into small blocks, after freehand section, placing the small blocks into distilled water in a container, performing lactic acid clearing, covering, dyeing via berberine or fluorescein, bleaching by the water, performing poststaining via toluidine blue O or sarranine O, bleaching by the water, performing water seal, observing and photographing via an inverted fluorescence microscope, and obtaining a clear image. The method can be used for observing the microscopic structures inside the plant roots, thereby being effectively used for the research on classification, auxanology, ecology and physiology of medical plants. The method disclosed by the invention has the advantages of simple method, easiness in operation, low cost, low working capacity, high success rate, and good effect, thereby being capable of being used for the research on morphological anatomy of the roots. The method is simpler and more effective for the thinner root sections of the plants.

The invention is about a method to purify the recombined human urokinase by the chromatography. The invention relates to recovery the gene engineering production from the mammal cell culture using the Streamline-SP, Sephacryl S-200, Sepharose Fast Flow and DEAE-Sepharose Fast Flow method. The recombined human urokinase can meet the SFDA standard and the percent recovery is above 70%, the purity is above 99% by using the method.

The invention relates to a method for producing batch hydrophobia vaccine, wherein it comprises that: (1), in the biological reactor with fixed bed mixing system and Fibra-Cel disks carrier, using cell increment cultivate liquid to cultivate Vero cell; (2), when the Vero cell grows to some density, using cell hold cultivate liquid, seeding hydrophobia vaccine, and affecting cell; (3), increasing virus; (4), obtaining virus continuously; (5), ultra-filter concentrating and inactivating via beta-propanolide; (6), using protamine sulfate or DNA enzyme treatment to remove Vero cell DNA; (7), using Sepharose 4 Fast Flow as stuff to process chromatography purification; (8), adding human blood albumin and sugar as protector to be made into liquid agent; or adding human blood albumin, sugar, and gelatin, as protector and shaping agent to be made into dried agent. The inventive method can produce virus continuously in small biological reactor, to realize batch production.

The invention relates to a method for preparing chloramphenicol affinant column and its usage, which adopts protein A -Sepharose 4 fast flow as phase carrier, the protein A of the carrier is coupled with the chloramphenicol antibody; the coupled chloramphenicol antibody is bonded with the fixed phase chemistry; the affinant column can suit for many stroma; it uses protein A so that IgG can be connected with it with high desirability and large load capacity of the chloramphenicol column.

The invention discloses single-decorative polietilenglicol insulin and making method with molecular weight at 6550-10800, which comprises the following steps: dissolving insulin in the acetate buffer with pH value at 3.0-5.0; adding mPEG-aldehyde and NaBH3CN; stirring; adding small-molecular amino acid to terminate reaction; dialyzing; condensing in the SP Sepharose FF chromatographic column; proceeding gradient elution through buffer with NaCl; collecting peak-3 eluent; condensing; dialyzing; freezing; drying to obtain PheB1-PEG-insulin.

The invention discloses a citrus fruit fly odorant binding protein-based attractant screening method belonging to the technical field of bioengineering. The citrus fruit fly odorant binding protein-based attractant screening method comprises the following steps of: collecting the total RNA (Ribonucleic Acid) of antennae of a citrus fruit fly; obtaining the overall length of a citrus fruit fly odorant binding protein through RT-PCR (Reverse Transcription-Polymerase Chain Reaction); constructing a prokaryotic expression vector of the citrus fruit fly odorant binding protein; inducing the expression of a citrus fruit fly recombinant odorant binding protein through IPTG (isopropyl-beta-d-thiogalactoside), and purifying the citrus fruit fly recombinant odorant binding protein through a nickel sepharose gel affinity column; obtaining a conjugation reaction spectrum of the citrus fruit fly recombinant odorant binding protein and a host fruit odor volatile matter through a competitive fluorescence combing method, wherein a dissociation constant (KD) is lower than below 10 mu mol/L host fruit smell; and the IC 50 value of fluorescence competition is less than 30 mu mol/L, determining a host fruit smell attractant suitable for the citrus fruit fly. The invention provides a new strategy for screening and designing a citrus fruit fly odorant host fruit smell odor information attractant formula.

The invention provides a method for preparing heparinase I. The method for preparing the heparinase I comprises the following steps of: inoculating flavobacterium heparinum serving as a raw material to a seed culture medium for culture; then inoculating the flavobacterium heparinum to a fermentation culture medium; centrifugally collecting precipitate; performing ultrasonication on the precipitate; performing centrifugation again to obtain crude enzyme liquid of the flavobacterium heparinum heparinase I; and performing SP-sepharose FF chromatographic purification on the crude enzyme liquid for three times to obtain the high-purity flavobacterium heparinum heparinase I, wherein the SP-sepharose FF chromatographic purification for three times is protected by calcium chloride in the whole course, so that the yield of pure enzymic activity is greatly increased. The method for preparing the heparinase I has the characteristics of simple process, easy amplification, large preparation amount of products at a time, low cost of reagents and the like. The specific activity of the prepared heparinase I reaches 223 IU/mg and the yield of the pure enzymic activity reaches 30 percent. Compared with the conventional newest method, the method for preparing the flavobacterium heparinum heparinase I has the advantages of increasing the specific activity by more than two times, and the purification yield by about one time.

The invention belongs to the technical field of deep wheat bran processing, and in particular relates to a method for extracting and purifying active polysaccharides of wheat bran. The method comprises the following steps: carrying out pretreatment, degreasing, polysaccharide extraction and crude polysaccharide refining; analyzing the yield, chemical composition and immunoregulatory activity of extracted crude products and purified components. By virtue of the method for extracting and purifying the active polysaccharides of the wheat bran, crude wheat bran polysaccharide products are prepared through a water extraction and alcohol precipitation method; the polysaccharides with the immunoregulatory activity are obtained by purifying a sepharose gel column. An extraction method is simple and the operation cost is low. The yield of the obtained polysaccharides of the wheat bran is 5.0 percent, the yield of the purified active polysaccharides of the wheat bran is 5.6 percent, and the polysaccharides have high immunoregulatory activity.

The invention discloses a grafting method for model arabidopsis thaliana.The method comprises the following steps that an arabidopsis thaliana plant with the seedling age of 6-8 days, thick and strong hypocotyl explant, consistent growth vigour and a main leaf emerging newly is used for grafting; stock and scion materials used for grafting are placed in the middle of 1.2% sepharose gel, the hypocotyl explant is transversely cut, and it is ensured that the hypocotyl explant part of the scion material is short and the hypocotyl explant part of the stock material is long; material notches are in tight contact on the sepharose gel, a 1/2MS+3% saccharose solid culture medium with the area larger than the stock root area is cut to cover the stock root, and a grafting body is formed; the grafting body is placed in the environment under the temperature of 21 DEG C to be vertically cultured for 7 days; after grafting notches are healed, the grafting body is transferred into a water culture device with 1/4MS as a liquid culture medium to be cultured.The problems that operation is difficult, notches are not round or normal, scion side roots are frequently produced, and the grafting survival rate is low due to the fact that arabidopsis thaliana plants are thin and weak are solved.

The invention belongs to the field of organic chemical synthesis, and relates to a method for connecting hyaluronic acid and sepharose. The method comprises the following steps: synthesizing hyaluronic acid-adipic acid dihydrazide derivatives by using the hyaluronic acid and adipic acid dihydrazide, activating the sepharose 4B by sodium periodate to obtain sepharose containing aldehyde group, and reducing the hyaluronic acid-adipic acid dihydrazide and the sepharose 4B containing the aldehyde group by sodium borohydride to obtain a connection substance of the hyaluronic acid and the sepharose 4B. The method has the advantages of mild reaction condition, simple steps and low cost.

The invention relates to a primer and method for identifying odontobutis potamophila, o. yaluensis and o. sinensis, particularly relates to identification of odontobutis potamophila, o. yaluensis and o. sinensis by applying microsatellite markers. A pair of primers disclosed by the invention is named as OP34a, and the sequences of the primers are shown in SEQ ID NO. 1 and SEQ ID NO. 2 ( as shown in the Specification). During the identification, the pair of the primers is used to perform PCR amplification on DNA samples of the three types of odontobutis obscura, and then 1.5% sepharose gel (dyed by EB) electrophoresis is used to detect PCR products; the products with 221-251 bp stripes are odontobutis potamophila, the products with 337-351 bp stripes are o. yaluensis, and the products with 465-471 bp stripes are o. sinensis. According to the invention, on the basis of without killing fingerlings, the identification can be performed by shearing a few tail fins or other fish tissue parts only, so that the primer and method have the advantages of high stability, simpleness in operation, excellent repeatability, high practicability, and rapid and accurate identification.

The invention discloses a heparin affinity column as well as the preparation method and the application thereof. The heparin affinity column is prepared through the following method: agarose gel 6FF is taken as a solid phase carrier which is firstly activated and then is aminated and coupled with amidocyanogen after an epoxy group is coupled, the aminated agarose gel 6FF and the heparin form intermediate aldimine in methanol, and the intermediate product of aldimine is further subjected to reductive amination for forming stable chemical bonds, thereby acquiring the heparin-agarose gel 6FF. The affinity column preparing preparation with high efficiency is simple and short. Through adopting the combination of the rear aldehyde of the heparin and the amidocyanogen of aminated sepharose, the activity of the heparin is not influenced, thereby ensuring the affinity column to have higher affinity; the heparin coupling is stable, and the heparin can be repeatedly used; the reagent adopted by the method has low cost and no environmental pollution, thereby laying a foundation for sweepingly isolating and purifying materials with specific binding capacity to the heparin in common laboratories, and promoting the research on domestic relative fields.

The invention provides a method for preparing sea cucumber fucoidan and sea cucumber glycoprotein. According to the method, acaudina molpadioides serves as the raw material, a mild stepped enzymolysis processing method is mainly adopted, an ultrafiltration membrane is utilized to separate out a mixture of crude polysaccharide and glycoprotein, then separation, purification and preparation of the sea cucumber fucoidan and the sea cucumber glycoprotein are achieved through a Q-Sepharose-F-F ion exchange column and an Sephadex G-150 sephadex column, a prepared sea cucumber fucoidan product is fawn, and the sea cucumber glycoprotein is faint yellow. According to the method, a co-production method is adopted, main ingredients such as polysaccharide and glycoprotein in the acaudina molpadioides can be prepared in a jointed mode, emission of waste is reduced in the extraction process, and production cost is lowered.

The invention discloses a nanogold solution preparing method based on sepharose gel. The method comprises the following steps that firstly, the sepharose gel is added into water and heated to obtain a clarification sepharose solution; secondly, a chloroauric acid solution is added into the sepharose solution and stirred evenly at the constant temperature, and the mixed solution is placed at the normal temperature to obtain the nanogold sepharose gel; thirdly, the nanogold sepharose gel obtained in the second step is dried in a freezing mode, the gel is removed, and the nanogold solution is obtained after washing. The nanogold solution is formed by evenly scattering nanogold particles in water, the sizes of the nanogold particles are even, and the average particle diameter ranges from 1 nm to 10 nm. The preparing technology is simple and practical, the applied reducing agent is the sepharose gel which is common and non-toxic, the sizes of the obtained nanogold particles are even and controllable, the grain diameter is small, and the nanogold particles are not prone to being aggregated. The nanogold solution preparing method has very important actual application value in the nanogold preparing field.

The invention discloses an acetylcholinesterase chemiluminescence bioreactor, and a preparation method and application thereof, and belongs to the technical fields of analysis of pesticide residues and biotechnology. The reactor is a glass tube of which the two ends are sealed by 300-mesh bolting silk, wherein immobilized acetylcholinesterase is held in the glass tube; and the immobilized acetylcholinesterase is purified crucian muscle acetylcholinesterase which is immobilized on cyanogen bromide-activated (CNBr) sepharose 4B. The application of the reactor is characterized by comprising the following steps of: establishing the acetylcholinesterase chemiluminescence bioreactor; and detecting organic phosphorus and carbamate type pesticides. The crucian muscle acetylcholinesterase (AChE) extracted in the invention has high enzyme activity, obvious purification effect and high immobilized enzyme activity recovery rate; the detection limit of the organic phosphorus type pesticides and the carbamate type pesticides is low; the analysis time is short; and the enzyme reactor can be repeatedly used.

The invention relates to primers and method for identifying greenhouse trialeurodes vaporariorum and bemisia tabaci by utilizing mitochondria SCAR (sequence characterized amplified regions) marker. The primers are nucleotide sequences shown as SEQ ID NO.1 and SEQ ID NO.2 respectively. The method comprises the following steps: (1) extracting aleyrodid genome DNA (deoxyribonucleic acid); (2) performing polymerase chain reaction (PCR) amplification by taking the aleyrodid genome DNA as a template; and (3) performing sepharose gel electrophoretic analysis on the PCR amplification prepared in the step (2). According to the invention, a simple and stable molecular marker for identifying the greenhouse trialeurodes vaporariorum and bemisia tabaci is provided and a foundation is laid for research of population dynamic identification, biology and invasion mechanisms of aleyrodid in the future.

The invention discloses a grifola frondosa mycelium anti-tumor glycoprotein and a preparation method, and belongs to the technical field of bioengineering. The glycoprotein is prepared by the following steps: using submerged fermented grifola frondosa mycelium as a raw material, homogenating and crushing, extracting with cold water, centrifuging, taking supernatant, precipitating with ammonium sulfate, dialyzing, and carrying out DEAE (diethyl-aminoethanol) -sepharose Fast Flow and superdexTM 75 prepgrad column chromatography and other steps to systematically separate and purify. The glycoprotein is a compound of polysaccharide and protein, wherein polysaccharide is 2-10% in content and composed of four monosaccharides including arabinose, fructose, mannose and glucose; the protein is 30-90% in content and composed of 17 amino acids including aspartic acid, methionine, glutamic acid and so on; the molecular weight of the glycoprotein is 30-90 KDa. The glycoprotein can inhibit growth of human gastric carcinoma cell SGC-7901 and human breast cancer cell MCF-7, and can be used for preparing a possible anti-cancer drug. Besides, the grifola frondosa mycelium anti-tumor glycoprotein can be generally applied to separation and purification of glycoprotein obtained from mycelium of various officinal and edible fungi through submerged fermentation.

The present invention discloses an anti-tumour snake venom cytotoxin and a production technique thereof, belonging to the fields of biochemistry and biological medicines. The snake venom cytotoxin of the present invention is derived from the prior lyophilized cobra venom powder. In the production technique of the present invention, the lyophilized cobra venom powder is resolved in the known buffer solution, and then on a SP-Sepharose H.P cation exchange column (5.0cm multipled by 29cm), gradiently eluted stage by stage by phosphate buffer and NaC1 at 10ml per tube and the flowing speed of 18ml per minute, a KTA Explorer protein purification system is used to monitor and collect eluting peaks, XII and XIII peaks with cytotoxic effects are collected, and via Source 30RPC reversed phase chromatography, cytotoxin XII and cytotoxin XIII, the molecular weight of which is about 6000 daltons to 7000 daltons and the isoelectric point of which is larger than 10, are respectively obtained. Compared with the purification step of the prior art, the production technique for producing the pure anti-tumour snake venom cytotoxin product without PLA2 has good rapid repeatability and is suitable for expanded production.

The present invention is characterized by that making the cysteine of 112th site of enterokinase light chain gene produce site-specific mutation, using colibacillus and microzyme to express enterokinase light chain variant, and utilizing Zn-Sepharose and STI-Sepharose affinity chromatography so as to obtain high-purity enterokinase light chain variant protein. As compared with wild enterokinase light chain said varient has higher stability and enzyme activity.

The invention relates to a primer and an identifying method for identifying different genetic collateral series of aphelinid. The identifying method comprises the following steps: (1), extracting aphelinid genome DNA (Deoxyribose Nucleic Acid); (2) carrying out PCR (Polymerase Chain Reaction) augmentation on a mitochondria COI (Cytochrome Oxidase Subunit) gene by adopting the aphelinid genome DNA as a template; (3), digesting the prepared PCR product by restriction enzyme Alu I to obtain enzyme-digested products; and (4), carrying out sepharose gel electrophoretic analysis on the prepared enzyme-digested products. The restriction enzyme is normal restriction enzyme, so that simple and stable enzyme digestion marks are provided for selecting the aphelinid with two genetic collateral series; and meanwhile, an identifying technology for the aphelinid with two genetic collateral series is established, so that the foundation is laid for population dynamics identification for the aphelinid with two genetic collateral series, the biology as well as the research of a mechanism of transmission.

The invention discloses a detection primer and a detection kit for watermelon wilt disease fungi. The detection primer is a dual-detection primer set consisting of a wilt disease fungus universal primer and a primer which only can amplify a watermelon wilt disease fungi. A reagent of the detection kit comprises the detection primer, a buffer solution, dNTPs, Taq polymerase, watermelon wilt disease fungus positive control DNA and ultrapure water. The invention further discloses a detection method. The detection method comprises the following steps: (1) extracting DNA of a plant tissue infected by the watermelon wilt disease fungi; (2) carrying out PCR amplification on DNA which is separated in the step (1) by virtue of the detection kit; and (3) separating PCR amplification products in the step (2) by virtue of sepharose gel electrophoresis, dyeing the products with ethidium bromide under an ultraviolet lamp after the separation, and determining a result according to the sizes of the amplified products. According to the invention, a method for detecting watermelon wilt disease fungi at a biological molecular level is provided and has the characteristics of high precision and convenient operation.

The invention discloses a method for increasing protein adsorption capacity and adsorption rate by using a high-density polyethylenimine (PEI) modified medium. The medium is a medium formed by modifying the PEI on the surfaces of sepharose gel particles with the average particle size of 50 to 170 Mum by an epoxy spacer arm. A preparation method comprises the steps of activating a chromatographic matrix by using epoxy chloropropane, and carrying out reaction between the activated medium and the amino group of the PEI to finish ligand coupling. The chromatographic medium has strong adsorption capability on protein in the range of 0.01 to 1 mol/L and shows high adsorption capacity and adsorption rate; the protein adsorption shows high salt concentration tolerance, so that material liquid can be directly contacted with the chromatographic medium, without being pretreated, to quick capture the target protein. The medium has a wide application prospect in high-efficiency and quick separation and purification of the protein.

The invention relates to a method for preparing a green tea polysaccharide through separation and purification and application of the polysaccharide in the preparation of anticoagulant health products and medicaments. The preparation method comprises the following steps: crushing green tea, leaching, performing alcohol precipitation, and performing vacuum freeze-drying to obtain a crude polysaccharide; performing dynamic decolorization and deproteinization through a polyamide column; and further purifying through a DEAE (diethylaminoethyl) sepharose CL-6B ion exchange column to obtain the green tea polysaccharide. The invention has the following technical effects: pigments and proteins in the green tea crude polysaccharide are effectively removed; and the components obtained through separation and purification and other raw materials and auxiliary materials which are acceptable in food and medicament processes can be developed into appropriate health food preparations and medicament products having an anticoagulant function, such as capsules, tablets, oral liquids and the like. Experiment proves that the polysaccharide is safe and non-toxic, has a favorable anticoagulant function in the preparation of anticoagulant health products and medicaments and has favorable development and application prospects.

The invention discloses a detection method of a folate metabolism related gene, which includes steps of extracting DNA from oral mucosa cell of a detector; amplifying MTHFR gene C677T by SEQ NO 1 and SEQ NO 2; amplifying MTHFR gene A1298C by SEQ NO 3 and SEQ NO 4; amplifying MTHFR gene A66G by SEQ NO 5 and SEQ NO 6; extracting DNA from oral mucosa cell as a template to perform PCR reaction; performing sepharose gel electrophoresis of 1% of ordinary PCR amplified product on three pairs of primers; recycling a strip of a kit recycling item from the 1% of agarose gel electrophoretic band by sepharose gel; performing sequence testing reaction; purifying the sequence testing reaction product and denaturing; testing sequence by an upper 3730 sequence tester; analyzing three SNP gene models of the sequence testing result by Chromas software. The method has the advantages of high detection accuracy, and convenience of use.

The invention relates to a deoxynivalenol immune affinity column and a preparation method and use thereof. The immune affinity purifying column is coupled to a sepharose supporter by protein G, and a deoxynivalenol antibody is coupled to the protein G on agarose; a crosslinking agent is used for crosslinking the deoxynivalenol antibody, the protein G and the sepharose supporter, and then the crosslinked deoxynivalenol antibody-protein G-sepharose supporter fills the affinity column. The immune affinity column is mainly used for purifying deoxynivalenol in food, feed, seasonings, wine, beverages and other various samples.