105results about How to "Improve the success rate of cultivation" patented technology

The invention discloses a culture medium for culturing fruiting bodies of Antrodia cinnamomea and a cultural method thereof. The culture medium comprises potato dextrose broth (PDB) with a concentration of 1-2%(w/v), glucose with a concentration of 2.8-3.6%(w/v), agar with a concentration of 2.2-4%(w/v) and a Chinese medicine composition with a concentration of 16.7-4.%(w/v). The cultural method comprises the following steps: preparing a solid culture medium according to the ratio of the ingredients, inoculating an Antrodia cinnamomea strain to the solid culture medium, putting the solid medium containing the Antrodia cinnamomea strain in a specific environment to culture for a certain period to generate the fruiting bodies of Antrodia cinnamomea, wherein the fruiting bodies of Antrodia cinnamomea obtained by the cultural method have same index components as wild Antrodia cinnamomea.

Provided is a plant tissue culture bacteriostatic (antiseptic) agent containing botanical fungicide. The bacteriostatic (antiseptic) agent comprises isothiazolinone, nisin and aqueous extracts of the Chinese herbal medicines of terminalia, groundsel and aromatic madder. When the bacteriostatic (antiseptic) agent is applied in plant tissue culture, 0.35 to 0.5% of the bacteriostatic (antiseptic) agent is added into a medium, which enables fungal and bacterial contamination to a culture to be effectively reduced or prevented and poses little adverse influence on plant growth, thereby enhancing the success rate of plant tissue culture; the bacteriostatic (antiseptic) agent can also be applied in open type plant culture, e.g., in the rooting phase of sound seedlings during a late plant culture stage, the bacteriostatic (antiseptic) agent is directly added into a medium, and the medium can be inoculated in indoor environment with bacteria or a few bacteria without undergoing the step of autoclaving or operating on a superclean bench, thereby simplifying seedling culture in plant tissue culture, enhancing working efficiency, saving a great amount of energy and reducing production cost for plant tissue culture seedlings; therefore, the bacteriostatic (antiseptic) agent has a critical actual application value.

A watermelon seed treatment method being high in germination rate comprises following steps: (1) pre-treatment of seeds: soaking watermelon seeds in an acidic soaking solution with flat seeds rejected; (2) preparation of a seed soaking liquid: mixing ammonium nitrate, sodium phthalate, sodium humate, an nutrient additive liquid and gibberellins to obtain a mixed aqueous solution; (3) seed soaking: soaking the pre-treated seeds in the seed soaking liquid; (4) sterilization treatment: treating the treated seeds under an infrared light lamp and sowing the seeds after sterilization. In the invention, the seeds are subjected to the pre-treatment so that dormant period of the watermelon seeds is broken, thereby promoting germination of the watermelon seeds. The seed soaking liquid is rich in nutrients so that the watermelon seeds can go through the dormant period with a short time, so that the speed of germination and seedling of the watermelon seeds is increased. The irradiation of the infrared light achieves the sterilization effect and can enhance disease resistance of the seeds, thereby increasing the cultivation successful rate of watermelon and achieving a high germination rate.

The invention belongs to the technical field of modern biotechnology cell culture. A purpose of the invention is to provide a tumor stem cell suspension culture method, which can be used for culture of tumor cells with different tissue sources, and tumor stem cells obtained through the culture method have characteristics of self-renewal capacity and enrichment. The tumor stem cell suspension culture method comprises: carrying out digestion centrifugation collection on tumor cells requiring culture, adopting PBS to carry out resuspending centrifugation, adding to a prepared suspension culture medium of the invention, carrying out beating-up into a single cell suspension, inoculating into a paved ultra-low absorption culture dish, and carrying out cell culture. The tumor stem cell suspension culture method has the following characteristics that: ball forming rates of various in vitro cultured tumor cells are high, multi-function factors are expressed, the OCT4 proportion and the NANOG proportion are increased, rich and comprehensive experimental materials are provided for tumor stem cell researches, and broad application prospects are provided in basic researches and clinical applications.

The invention discloses a method for culturing bladder cancer organs in vitro. The method comprises the following steps: manufacturing a gas-liquid interaction culture system, namely manufacturing a uniformly paved rat tail collagen supporting layer on the surface of a porous culture membrane in a Transwell upper chamber; re-suspending fresh in-vitro bladder cancer tissues by using a rat tail collagen solution, uniformly mixing, adding the obtained mixture onto the rat tail collagen supporting layer, and then putting the rat tail collagen supporting layer into an incubator at 37 DEG C to solidify so as to obtain a rat tail collagen layer containing the bladder cancer tissues; adding an organoid culture medium into a Transwell lower chamber, and enabling the liquid level of the culture medium to be lower than the rat tail collagen layer containing bladder cancer tissues; and carrying out passage and cryopreservation. The culture success rate of the bladder cancer organoid is remarkablyincreased, the bladder cancer organoid with reserved immune cells can be obtained through culture, operation is easy, the utilization rate of tumor tissue is high, and great significance and value areachieved for bladder cancer drug screening research.

The invention puts forward a universal cancer organoid in-vitro culture medium, which consists of a DMEM/F-12K (Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12K) basic culture medium and an adding factor, wherein the adding factor consists of HEPES (2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid), L-Glutamine, EGF (epidermal growth factor), Noggin, FGF (fibroblast growth factor)-10, A83-01 and Y27632. According to the culture medium of the embodiment of the invention, the success rate of organoid passage culture can be greatly improved, in addition, the universal culture of cancer samples, including gastric carcinoma, rectal carcinoma, lung carcinoma and the like, can be realized, culture cost is greatly lowered, the long-term culture of the organoid can be realized, and abiobank is established.

The preparation process of cell-eliminating pig corium substrate includes sustained vibration of the corium substrate in 0.5 mol/L NaOH solution compounded with thrice distilled water, rinsing with thrice distilled water and balanced phosphate salt solution separately, freeze drying, packing and sterilizing. The cell-eliminating pig corium substrate is used as the in vitro corium culturing rack in composite skin preparing tissue engineering process, and the preparation process includes coating the corium substrate with acetic acid dissolved type-I ox collagen, regulating pH to 7.0-7.4, soaking subsequently in balanced phosphate salt solution and no serum culture medium, inoculating passage epidermal cell, and no serum culture inside a CO2 hatcher. The tissue engineering composite skin is used as human skin substitute.

The invention discloses a method of cultivating Gastrodia elata and belongs to the technical field of planting of Gastrodia elata. The method comprises the steps of preparing Armillaria mellea, preparing mushroom sticks, inoculating, cultivating Gastrodia elata, and managing. By controlling the cultivation process of Armillaria mellea and modifying the method of cultivating Gastrodia elata, the yield of Gastrodia elata is increased; tubers of Gastrodia elata cultivated through the method are uniform in size, full and good in condition.

The disclosure relates to a method for culture of a cis-platinum drug-resistant lung cancer organoid. The method comprises the following steps: mixing cis-platinum drug-resistant lung cancer tissue cells with the medium and matrix gel to obtain a to-be-cultured substance, wherein the medium contains matrix metalloproteinase-10, and the concentration of the matrix metalloproteinase-10 is 50-200nM based on the medium; and carrying out culture amplification on the to-be-cultured substance to obtain the cis-platinum drug-resistant lung cancer organoid. According to the culture method for the cis-platinum drug-resistant lung cancer organoid provided by the invention, since the employed medium contains the matrix metalloproteinase-10 with specific concentration, and the matrix metalloproteinase-10 with the specific concentration can promote in-vitro growth of the cis-platinum drug-resistant lung cancer organoid, the success rate of the culture is high when the method provided by the disclosure is used for the culture of the cis-platinum drug-resistant lung cancer organoid.

The invention relates to a method for improving the success rate of umbilical cord blood mesenchymal stem cell culture by using placental mesenchymal stem cells. The method comprises the following steps: firstly, digesting placentallobules in a short time by using collagenase I to obtain a paste mixture consisting of extracellular matrix protein and tissue blocks and carrying out primary culture; and then co-culturing third-generation placental mesenchymal stem cells as moisturizing cells and separated primary umbilical cord blood mesenchymal stem cells under a low-oxygen condition to obtain a lot of umbilical cord blood mesenchymal stem cells. The method for improving the success rate of umbilical cord blood mesenchymal stem cell culture by using placental mesenchymal stem cells provided by the invention shortens the culture time of the placental mesenchymal stem cells, effectively solves the problem that the placental mesenchymal stem cells are low in purity and poor in multiplication capacity and the like, improves the success rate of umbilical cord blood mesenchymal stem cell culture and develops the utilization value of the umbilical cord blood mesenchymal stem cells. As fetal calf serums and pancreatin are not used, the safety of the cells on clinical application is enhanced.

The invention relates to a method for culturing liver cancer organoid. The method comprises the following steps: mixing liver cancer tissue cells with a culture medium and matrix gel to obtain a to-be-cultured substance, wherein the culture medium containing an insulin growth factor-2, and the concentration of the insulin growth factor 2 being 1-50ng/ml, preferably 5-10ng/ml based on the culture medium; and carrying out culture amplification on the to-be-cultured substance to obtain the liver cancer organoid. According to the culture method of the liver cancer organoid provided by the invention, the adopted culture medium contains the insulin growth factor-2 with the specific concentration, and the insulin growth factor-2 with the specific concentration can promotes in-vitro growth of theliver cancer organoid, so that the culture success rate is relatively high when the method provided by the invention is used for culturing the liver cancer organoid.

The invention relates to a method for culturing a dedifferentiated or undifferentiated thyroid carcinoma organ. The method comprises the following steps: mixing dedifferentiated or undifferentiated thyroid carcinoma tissue cells with a thyroid carcinoma culture medium and matrix glue to obtain a to-be-cultured substance, wherein the thyroid carcinoma culture medium contains nicotinamide and BM-Cyclin, and based on the thyroid carcinoma culture medium, the concentration of nicotinamide is 5-20 mM, and the concentration of BM-Cyclin is 5-30 [mu]g/ml; and carrying out culture amplification on theto-be-cultured substance to obtain the dedifferentiated or undifferentiated thyroid carcinoma organ. In the method for culturing the dedifferentiated or undifferentiated thyroid carcinoma organ, theadopted thyroid carcinoma culture medium contains nicotinamide and BM-Cyclin with specific concentrations, so that the in-vitro growth of the dedifferentiated or undifferentiated thyroid carcinoma organ is promoted. Therefore, when the method provided by the invention is used for culturing the dedifferentiated or undifferentiated thyroid carcinoma organ, the culture success rate is relatively high.

The invention relates to a biomedical technology, in particular to a culture medium and culture method for ovarian cancer organoids. The culture medium comprises a basal culture medium DMEM/F12, B27, glutamine, HEPES, N-acetylcysteine, FGF2, A83-01, nicotinamide, forskolin, beta-estradiol, hydrocortisone, R-spondin-1, noggin, FGF10, EGF and Heregulin beta 1 and at least one selected from PFBS, PFOA and PFOS. According to the culture medium and the culture method, the establishment success rate of an ovarian cancer culture medium is improved, the amplification efficiency of the ovarian cancer organoids is improved, the proliferation efficiency after passage is stable, the drug screening process is accelerated, and the cost of culture materials is reduced.

The invention discloses a culture method of human colorectal cancer tissue organoid, which comprises the following steps: washing human colorectal cancer tissues and putting them into a transfer culture medium, cleaning and shearing; further cleaning, centrifuging, carrying out enzyme digestion, filtering, and collecting cell clusters; and mixing the cell clusters with hydrogel, adding DMEM culture medium to culture after solidification to obtain the human colorectal cancer tissue organoids. According to the invention, gentamicin, amphotericin B, primocin and other antibiotics are added into the transfer culture medium, the digestion solution and the culture medium, so that microbial contamination can be reduced, and the method for obtaining the human colorectal cancer organoids through in-vitro culture is simple, convenient, rapid, high in success rate and good in organoid growth state. The invention further discloses application of the human colorectal cancer organoid in drug testing, and the co-culture model of human colorectal cancer tissue organoids and tumor-associated fibroblasts can be used as a patient individualized drug testing platform.

The invention relates to a method for preparing an ovarian cancer organoid, which is characterized by comprising the following steps of: a, mixing feeder layer cells, temperature-sensitive hydrogel, ovarian cancer cells and a first culture medium, and culturing for 7-9 days to obtain a first culture; wherein the feeder layer cells are human endometrial stem cells without a proliferation function;and b, changing the liquid of the first culture by using the first culture medium, and culturing until the ovarian cancer organoid is formed. According to the method disclosed by the invention, the ovarian cancer organoid matched with ovarian cancer tissue biological information can be cultured by utilizing a small amount of ovarian cancer tissues or cells, and the culture success rate is relatively high.

The invention relates to the technical field of cell preservation and transportation, and particularly discloses a low-temperature preservation solution and a preservation method thereof. The low-temperature preservation solution is prepared from a DMEM high-glucose culture medium, Glutamax, HEPES and Penicilline-Streptomycin. The organoids transported by the cryopreservation solution at low temperature do not need to be resuscitated, and can be quickly cultured, is relatively low in cost and relatively high in culture success rate, and the activity of cells can be maintained for a long time. According to the invention, the technical problems of high hardware requirements, complex transportation procedures, high cost, low efficiency, low success rate of cryopreservation recovery, worry about cell growth conditions and the like are solved.

The invention relates to a full-automatic multifunctional cell treatment system which comprises a biosafety cabinet and a workbench, and the workbench comprises an on-table part and an off-table part; wherein the on-table part of the workbench comprises a balancing unit, a heating unit, a quality control unit, a connection unit, a sample unit, a reagent unit, a consumable unit, a gun head unit, a waste unit, a liquid suction device, a first controller, a cap screwing device with recognition and clamping functions, and a second controller; the off-table part of the workbench comprises a centrifugal machine and a power supply control module. The biosafety cabinet is arranged outside the workbench, and a switch window is arranged on the biosafety cabinet.

The invention discloses a method for cultivating gastrodia tuber by clinker wood. The method comprises the following steps: taking the raw materials of 90-110 parts of raw material saw dust, 50-70 parts of wheat bran, 15-25 parts of corn flour, 1-3 parts of brown sugar and 1-3 parts of plaster powder; evenly mixing the raw materials to obtain a mixed filler for backup; segmenting the wood into 15cm-small-wood sections; binding the 15cm-small-wood sections and then loading into a strain bag, filling up gaps in the strain bag with the obtained mixed filler, and then fastening the opening of the bag; placing the bound strain bag into a sterilizing pot for sterilization; heating to 100 DEG C under the condition of normal-pressure sterilization and then keeping for 8-10 hours; after stopping fire, keeping for 2 hours, and then opening an oven door for heat dissipation; cooling the temperature of the bag to 70 DEG C, getting the bag out of an oven, and then transferring the bag into a cooling room; and finally cooling, inoculating, culturing the wood indoors, and cultivating outdoors. The method of the invention carries out high-temperature disinfection treatment on the wood, thus achieving less mixed wood pollution, short wood cultivation time, high culture success ratio of the wood and hypha and saved resources.

The invention discloses a primary culture method of the embryonic cell tissue pieces of a Chinese softshell turtle. The primary culture method comprises the following steps of: (1) collecting the fertile eggs of the Chinese softshell turtle, and disinfecting; (2) breaking the eggshells of the fertile eggs, separating embryos from yolk, cleaning the separated embryonic tissues with a 1*PBS (Phosphate Buffer Solution), and then soaking for sterilization; (3) cleaning the embryonic tissues for 4-5 times in an M199 culture medium, then transferring into a complete culture medium, cutting the embryonic tissues into pieces to obtain tissue pieces, and then cleaning the tissue pieces once by using the complete culture medium; (4) uniformly dispersing the tissue pieces to the inner wall of a cell bottle by using a sterile disposable inoculating needle so that the tissue pieces grow for 3-4 hours in a way of being attached to the wall, then adding the complete culture medium to submerge the tissue pieces, and culturing at the constant temperature of 30-31 DEG C for more than 3 days. The primary culture method disclosed by the invention has the advantages of easy operation, convenient culture, no need of specific cell separation liquid and high culture success rate.

The invention provides a three-dimensional sphere-gathering culture chamber mold and a method of screening concentrations of chemotherapeutic drugs in cancer cells, wherein the three-dimensional sphere-gathering culture chamber mold comprises at least one mold post for making cell culture holes via casting; each mold post a first segment, a second segment and a semisphere connected coaxially in sequence in a casting insertion direction, the axial length of the first segment is 20-30 times greater than the radius of the semisphere, the axial length of the second segment is 10-15 times greater than the radius of the semisphere, the second segment includes at least two buffer segments stacked, the sidewall of each buffer segment is inclined to the axis of the mold post in the insertion direction, and the inclination angles are different. The three-dimensional sphere-gathering culture chamber mold provided herein is suitable for making cell culture holes to simulate the human environment on an existing orifice plate through casting, so that efficient three-dimensional cell culture is achieved. In addition, the buffer segments enable the cell culture holes to correspondingly form a buffer chamber with multilayer buffer structure, so that cell damage caused during liquid changing is avoided.