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微信小章| 产品名称： | Dermamoeba minor (Pussard et al.) Page |
|---|---|
| 商品货号： | TS182962 |
| Strain Designations： | GMU-1 |
| Biosafety Level： | 1
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country. |
| Isolation： | pond sediment, George Mason University, Manassas, VA, 2001 |
| Product Format： | frozen |
| Type Strain： | no |
| Medium： | ATCC® Medium 919: Non-nutrient agar |
| Growth Conditions： | Temperature: 25.0°C Protocol: Both TS182962 and the food organism, Chlorella sp. ATCC 30562, are shipped frozen. Upon their arrival, establish a culture of ATCC 30562 as soon as possible. It is best to order ATCC 30562 in advance of ordering TS182962 unless you have liquid nitrogen storage facilities. When ATCC 30562 arrives, place it directly into a 35C-water bath and transfer its thawed contents to a fresh plate of ATCC medium 5. Incubate the plate at 25C under a 14/10-hour light cycle under 50-100 microEinsteins/meter squared per second of irradiance. When there is a dense lawn of cells, suspend the cells using ATCC medium 1323 or some other balanced salt solution. Transfer a 0.2-ml aliquot of a dense suspension of the alga to an agar plate of ATCC medium 919 and spread it evenly over the surface with a spread bar. Thaw the frozen ampule of TS182962 in a 35C-water bath. Transfer the entire contents of the vial to the agar plate of ATCC medium 919 with the lawn of Chlorella. Incubate the plate upright in the dark at 25C. Maintain cultures of the food organism on ATCC medium 5 as described above. |
| Subcultivation： | Protocol: Both TS182962 and the food organism, Chlorella sp. ATCC 30562, are shipped frozen. Upon their arrival, establish a culture of ATCC 30562 as soon as possible. It is best to order ATCC 30562 in advance of ordering TS182962 unless you have liquid nitrogen storage facilities. When ATCC 30562 arrives, place it directly into a 35C-water bath and transfer its thawed contents to a fresh plate of ATCC medium 5. Incubate the plate at 25C under a 14/10-hour light cycle under 50-100 microEinsteins/meter squared per second of irradiance. When there is a dense lawn of cells, suspend the cells using ATCC medium 1323 or some other balanced salt solution. Transfer a 0.2-ml aliquot of a dense suspension of the alga to an agar plate of ATCC medium 919 and spread it evenly over the surface with a spread bar. Thaw the frozen ampule of TS182962 in a 35C-water bath. Transfer the entire contents of the vial to the agar plate of ATCC medium 919 with the lawn of Chlorella. Incubate the plate upright in the dark at 25C. Maintain cultures of the food organism on ATCC medium 5 as described above. |
| Cryopreservation： | 1.xa0xa0 Harvest cells from a culture which is at or near peak density by adding 3.0 ml fresh ATCC medium 5 broth to the slant and washing cells into suspension. 2.xa0xa0 Adjust the concentration of cells to 2 x 106 -2 x 107xa0 / ml with fresh broth medium, then dilute to half this concentration by adding an equal amount of a 10% (v/v) sterile methanol solution in fresh ATCC medium 5 broth. 3.xa0 Dispense in 0.5 ml aliquots into 1.0 - 2.0 ml sterile plastic screw-capped cryules (special plastic vials for cryopreservation). The time from the mixing of the cell preparation and methanol stock solution to the beginning of the freezing process should be no less than 5 min and no greater than 15 min. 4.xa0xa0 Place vials in a controlled rate freezing unit. From room temperature cool at -1°C/min to -40°C. If freezing unit can compensate for the heat of fusion, maintain rate at -1 C/min through heat of fusion. At -40°C plunge ampules into liquid nitrogen. Alternatively, place the vials in a Nalgene 1°C freezing apparatus.xa0 Place the apparatus at -80°C for 1.5 to 2 hours and then plunge ampules into liquid nitrogen.xa0 (The cooling rate in this apparatus is approximately -1°C/min.) xa0xa0 5.xa0xa0 The frozen preparations should be stored in either the vapor or liquid phase of a nitrogen refrigerator. Frozen preparations stored below -130°C are stabile indefinitely. Those stored at temperatures above -130°C are progressively less stabile as the storage temperature is elevated. Vials can be stored between -80 and -70°C for no longer than one week. 6.xa0xa0 To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial to a level just above the surface of the frozen material. Do not agitate the vial. 7.xa0xa0 Immediately after thawing, do not leave in the water bath, aseptically remove the contents of the ampule and add to a centrifuge tube containing 5 ml of ATCC medium 5 without agar.xa0 Centrifuge at 300 x g for 5 min. 10.Remove most of the supernatant (=methanol, which can inhibit growth) and then resuspend the pellet.xa0 Transfer the culture to a fresh tube of ATCC medium 5 and incubate on a 15° horizontal slant at 50-100 µEinsteins/m2/s irradiance at 25°C with the cap loosened one half turn.xa0 Maintain under a 14/10 h light-dark photoperiod and subculture every 14-21 days. |
| Name of Depositor： | TA Nerad |
| Special Collection： | NSF - Protistology |
| Year of Origin： | 2001 |
| References： | Thomas A Nerad, personal communication |