TH10745C3 - The process of preparing invasive cell suspensions to produce glucomannan - Google Patents

Abstract

DC60 (14/09/15) Preparation method for invasive cell suspension using the culture of suspended cell suspension under An appropriate control condition with procedures consisting of Preparation of aseptic konjac tissue, inducing sterile konjac tissue, formation and development, etc. Inducing a fragment of invasion callus creating a fireball callus And suspended cell culture invade the fibrous callus To get cells to invade at Is stable in storage and transplantation. For use as an appropriate starting cell in production Glucomannan in bioreactor Revised Conclusion 06/03/2015 Procedure for preparing invasive cell suspensions by using suspended cell culture technique under An appropriate control condition with procedures consisting of Preparation of aseptic konjac tissue, inducing sterile konjac tissue, formation and development, etc. Induction of sterile konjac fragments to callus formation. And suspended cell culture invade the fibrous callus To get cells to invade at Is stable in storage and transplantation. For use as an appropriate starting cell in production Gluconumannan in bioreactor ------------------------------------------------ Edit 14 / 09/2015 Method for preparing invasive cell suspensions by using the culture technique of suspended cell suspension under An appropriate control condition with procedures consisting of Preparation of safe konjac tissue, inducing sterile konjac tissue, formation and development, etc. Inducing a fragment of konjac callus creating a friable callus. And suspended cell culture invade the fibrous callus To get cells to invade at Is stable in storage and transplantation. For use as an appropriate starting cell in production Gluconumannan in bioreactor -------------------------------------------------- ---------------------------- Preparation method for invasive cell suspension using culture technique An appropriate control condition with procedures consisting of Preparation of safe konjac tissue, inducing sterile konjac tissue, formation and development, etc. Inducing a fragment of invasive callus creating a fireball callus. And suspended cell culture invade the fibrous callus To get cells to invade at Is stable in storage and transplantation. For use as an appropriate starting cell in production Gluconumannan in bioreactor

Claims (5)

Disclaimer (all) which will not appear on the advertisement page: Amendments 6/3/2015 1. The method for preparing invasive cells to produce glucomannan The steps are as follows: a. Preparation of sterile konjac tissue from the shoots and head on the leaves in an amount of 20-50 g per 100 mm bleach solution by immersing in a 70% (vol / volume) alcohol solution and bleaching. Sterilized by 20% sodium hypochlorite (NaOCl) solution (volume / volume) for 20 and 25 minutes respectively, then cultured on solid base MS formula with 5.8 pH. The incubation was performed under 16 hours of light per day and temperature of 25-27 ° C for 7 days, as in Example 1 b. Induction of sterile konjac tissue to form shoots and multiply the head parts on the leaves and the plant. Konjac shoots by using 2-10 grams of sterile konjac tissue to cultivate on Solid MS formula has a pH of 5.8 with a growth regulator. 6- Benzyladenine (6- Benzyladenine; BA), concentration 2.0 mg / l. By incubation under 16 hours of light per day and temperature of 25-27 ° C for 30-45 days, the completely sterile shoots of the konjac were cut 2-10 g Is it possible to have the volume?) Cultured on the basic solid medium of MS formula with the pH 5.8 with the BA growth regulator at the concentration of 0.5 - 2.5 mg / l. Curing at 25? 2 ° C under lighting conditions for 16 hours per day for 60 days to multiply the peaks. For example 2, Fig. 2 c. Induction of parts Of sterile konjac plant calluses and callus proliferation caused by Parts of the leaves, stems and roots of the parts. On basic solid MS formula with pH 5.8 with growth regulator 2,4-Dichlorophenoxy acetic acid (2,4-Dichlorophenoxy acetic acid; 2,4-D) by murmuring under 16-hour light / day or no-light conditions, temperature. 25? 2 ° C for 60 days and applied to induce Fireable Callus For example 3 d. The increase of callus content from callus prepared from fragments of stem and stem leaves by cutting the parts to cultivate on solid MS formula with high strength. It was 5.8-pH with the growth regulator 2,4-dichlorophenoxyacetic acid by incubation under 16 h / day exposure to temperature 25-27 ° C for 60. Example 4: Induction of callus into fry calluses by dividing the callus prepared from the And the appropriate recipes above About 10 mm in diameter. Basic solid MS formula with growth regulators 2,4-dichlorophenoxyacetic acid was incubated under 16 h / day exposure at 25? 2 ° C for 60 days as example 5. Calluses are formed from sterile fragments of the konjac leaves, stems, and bases. It was cultured on basic solid MS formula with pH 5.8 with growth regulators, naphthalene acetic acid (NAA). There are 16 hours of light per day for 30 days to induce calluses and increase the number of calluses to use. In inducing fibrous calluses For example 6 f. Increase the amount of fibrous callus by dividing 1 - 2 grams of the fibrous callus prepared from d. To cultivate on solid MS formula. With growth regulator 2,4-dichlor The rophhenoxacetic acid was incubated under the light conditions at 25-27 ° C for 60 days. Example 7 h. The 250 ml flask was cultured in a basic liquid diet of MS formula with pH 5.8 with 2,4-D growth regulator and 50 ml functional sucrose. Incubated on a vibrator at a speed of 120 rpm, temperature 25? 2 degrees Celsius, changed the food every 7 days for 60 days as shown in Example 8 2. The preparation of invasive cell suspension to produce glucomannan According to claim 1, where optimum conditions are 16 hours of exposure per day, temperature 25? 2 degrees Celsius. 3. Preparation of invasive cell suspension to produce glucomannan. According to claims 1-2, any one that, in the process of inducing various parts Of the sterile konjac produces that callus The growth regulators used are selectable from In any case, 2,4-dichlorophenoxyacetic acid and / or naphthalene acetic acid (NAA) in any way. 4. Preparation of invasive cell suspension for production. Glucomannan According to one of the claims 1-3, where in the culture of invasive cell suspension The MS formula liquid food used may contain 0.5 and / or 0.75 mg / l of 2,4-dichlorophenoxy acid. 5. The preparation of invasive cell suspension to produce glucomannan. A According to any one of the rights 1-4 that in the culture of invasive cell suspension Can be cultured in Or without light -------------------------------------------------- ------ Revised 14/09/2015 1. The preparation of invasive cell suspension to produce glucomannan The steps are as follows: A. Preparation of sterile konjac tissue from the shoots and heads on the leaves in an amount of 20-50 g per 100 ml of bleaching solution by soaking in a 70% alkaline solution (volume It was then sterilized with 20% sodium hyperchlorite (NaOCL) solution (volume / volume) for 20 and 25 minutes respectively. The pH 5.8 was incubated under 16 hours of photocatalytic conditions and temperature of 25-27 ° C for 7 days. B. Induction of sterile konjac tissue to form peaks and multiplication of tubers. The leaves and shoots of konjac by using 2-10 grams of konjac pulp that were sterilized and treated on Solid MS formula has a pH of 5.8 with a growth regulator. 6- Benzyladenine (6- Benzyladenine; BA), concentration 2.0 mg / l. It was incubated under 16 hours of light per day and temperature of 25-27 ° C for 30-45 days. Completely sterile shoots of 2-10 g of konjac were cultured on solid food basis. The MS formula with pH 5.8 with the BA growth regulator was incubated at 25? 2 ° C under 16 h / day light conditions for 60 days to increase the multiplicity. Different parts Of sterile konjac plant calluses and callus proliferation caused by Parts of the leaves, stems and roots, especially the parts. On basic solid MS formula with pH 5.8 with growth regulator 2,4-Dichlorophenoxy acetic acid (2,4-Dichlorophenoxy acetic acid; 2,4-D) was cured under 16-hour exposure per day or no-light condition. 25 + 2 ° C for 60 days and applied to induce Fine Callus For example 3 d. The increase in the amount of callus from callus obtained from parts of the stem and root by cutting the parts to cultivate on solid MS formula that The pH 5.8 with the growth regulator 2,4-dichlorophenoxyacetic acid was incubated under 16 h / day exposure to temperature 25-27 ° C. 60 days e. Induction of calluses to become fry calluses by dividing the prepared calluses from the parts. And the appropriate solid food recipes above About 10 mm in diameter, cultured on food Basic solid MS formulation with growth regulator. It was then cured under 16 h / day exposure at 25? 2 ° C for 60 days. Calluses may also be induced. The fragments of the leaves, stems and bases of sterile konjac were cultured on solid MS formula with pH 5.8 with naphthalene acetic growth regulator (Napthalene). Acetic Acid; NAA) by incubation under 16 hours photoday for 30 days to induce calluses and increase the number of calluses to be used. To induce the friable callus f. Increase the amount of fibrous callus by dividing 1-2 grams of the fibrous callus prepared from the D. It was cultured on solid MS formula containing 2,4-dichlorophenoxyacetic acid by incubation under light conditions at 25-27 ° C as 60 days h. Cell culture of konjac suspension from fibrous callus in 250 ml flask was cultured in basic liquid diet, MS formula with pH 5.8 with 2,4-D growth regulator and 50 ml working volume sucrose by grumbling on shaker at 120 rpm, temperature 25? 2 ° C, changing food every 7 days for 60 days 2. The process of preparing invasive cell suspensions to produce glucomannan According to claim 1, where optimum conditions are 16 hours exposure per loop temperature 25? 2 degrees Celsius. 3. Preparation of invasive cell suspension to produce glucomannan. According to claims 1-2, any one that in the process of inducing the parts Of the sterile konjac produces that callus The growth regulators used are selectable from 4. The preparation of invasive cell suspensions to Production of glucomannan According to one of the claims 1-3, where in the culture of invasive cell suspension The MS formula liquid food used may contain 0.5 and / or 0.75 mg / l of 2,4-dichlorophenyrene acid. A According to any one of the rights 1-4 that in the culture of invasive cell suspension Can be cultured in Or without light ------------------------------------------------- 1. The process of preparing invasive cells to produce glucomannan The steps are as follows: A. Preparation of sterile konjac tissue from the shoots and heads on the leaves in an amount of 20-50 g per 100 ml of bleaching solution by soaking in a 70% alkaline solution (volume It was then sterilized with 20% sodium hyperchlorite (NaOCL) solution (volume / volume) for 20 and 25 minutes respectively. The pH 5.8 was incubated under 16 hours of exposure per day and temperature 25-27 ° C for 7 days, as in Example 1 b. The head parts on leaves and shoots of konjac were carried out by using 2-10 grams of konjac pulp that was sterilized and treated on Solid MS formula has a pH of 5.8 with a growth regulator. 6- Benzyladenine (6- Benzyladenine; BA), concentration 2.0 mg / l. It was incubated under 16 hours of light per day and temperature of 25-27 Celsius for 30-45 days. How much is the volume?) Cultured on the MS formula basic solid food with 5.8 pH with the growth regulator BA at the concentration of 0.5-2.5 ml / g. Curing at 25 + 2 ° C under lighting conditions for 16 hours per day for 60 days in order to multiply. For example 2, Fig. 2 c. Induction of parts Of sterile konjac plant calluses and callus proliferation caused by Parts of the leaves, stems and roots, especially the parts. On basic solid MS formula with pH 5.8 with growth regulator 2,4-Dichlorophenoxy acetic acid (2,4-Dichlorophenoxy acetic acid; 2,4-D) was cured under 16-hour exposure per day or no-light condition. 25 + 2 ° C for 60 days and applied to induce Fine Callus For example 3 d. The increase in the amount of callus from callus obtained from parts of the stem and root by cutting the parts to cultivate on solid MS formula that The pH 5.8 with the growth regulator 2,4-dichlorophenoxyacetic acid was incubated under 16 h / day exposure to temperature 25-27 ° C. Time 60 days and apply induction Fireable Callus For example 3 e. Inducing callus into fry callus by dividing the callus prepared from the part And the appropriate solid food recipes above About 10 mm in diameter. Basic solid MS formula with growth regulators It was then cured under 16 h / day exposure at 25 + 2 ° C for 60 days as example 5. Calluses are formed from sterile fragments of konjac leaves, stems, and bases. In particular, it was cultured on basic solid MS formula with pH 5.8 with naphthalene acetic growth regulator (NAA). 16 hours of light for 30 days to induce calluses and increase the number of calories used. In inducing fibrous calluses For example 6 f. Increase the amount of fibrous callus by dividing 1-2 g of fibrous callus prepared from section D to cultivate on solid MS formula. There were 2,4-dichlorophenoxyacetic acid regulators by incubation under the light condition at 25-27 ° C for 60 days. 7 g. Cell culture suspension of konjac friable callus in 2501 ml flask was cultured in basic liquid MS formula with pH 5.8 with a controlled substance. The growth of 2,4-D and 50 ml working volume sucrose by complaining on a shaker at 120 rpm, temperature 25 + 2 ° C, changing food every 7 days for 60 days, as in example 8. 2. The process of preparing invasive cells to produce glucomannan According to claim 1, where the optimum condition is 16 hours exposure per loop temperature 25 + 2 degrees Celsius. 3. The process of preparing invasive cells to produce glucomannan According to claims 1-2, any one that, in the process of inducing various parts Of the sterile konjac produces that callus The growth regulators used are selectable from Either 2,4-dichlorophenoxyacetic acid and / or naphthalene acetic acid (NAA). 4. Method for preparing invasive cell suspension to produce glucomannan According to one of the claims 1-3, where in the culture of invasive cell suspension The MS-based liquid diets may contain 0.5 and / or 0.75 mg / l of 2,4-dichlorophenoxyacetate. 5. Method for preparing invasive cell suspension to produce glucomannan According to any one of the rights 1-4 that in the culture of invasive cell suspension Can be cultured in Or without light