in a clean, non-sterile 15 mL centrifuge tube, prepare a 2.0% yeast suspension by adding 0.06 g Saccharomyces cerevisiae to 3 mL yeast growing medium (56 mM glucose, 20 mM HEPES, pH 6.8). What percent of yeast suspension is left after a 1:10 dilution?
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in a clean, non-sterile 15 mL centrifuge tube, prepare a 2.0% yeast suspension by adding 0.06 g Saccharomyces cerevisiae to 3 mL yeast growing medium (56 mM glucose, 20 mM HEPES, pH 6.8).
What percent of yeast suspension is left after a 1:10 dilution?
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- Imagine you have been given a liquid culture of yeast with a starting concentration of 3.67 x 10' cells/ml and are asked to carry out the sample dilution process shown in the figure below. 100μl 100μl 100μl 100μl 100μl 0.9ml 0.9ml 0.9ml H2O H₂O 6.9ml 0.9ml H₂O H₂O H₂O Original 10-1 102 10-3 104 Culture 105 100μl 100μl 100μl Plate A Plate B Plate C a. How many colonies should have been present on Plate A in this example? - Answers must be whole numbers as partial colonies are not expected. b. Imagine you carried out the same dilution scheme shown in the figure above, but now, you do not know the concentration of the original culture. If you counted 163 colonies on Plate B, what is the concentration of cells/ml in the original culture?A culture of S. cerevisea has an overnight OD of 4.5 (1.0 OD is approx 1.0x107 cells/ml) You will be plating 100µl onto agar and want the final count of colonies on the plate to be around 300 colonies. How much of the 4.5 OD culture must you use to get a 500µl subdilution (with sterile water), so that you have diluted enough to get approx 300 colonies per 100ulInoculate 250-µL overnight cell culture into 50 ml LB medium (in a 250 ml flask). Shake vigorously at 37 °C to OD600~0.5-0.6 (usually it takes about 2-3 hours). Why should we use a larger flask during culture at this step? Why do we need to wait till this OD range is achieved?
- Two flasks of E. coli are grown in batch culture in the same medium (2% glucose and amino acids; no nitrate) and at the same temperature (378C). Culture #1 is well aerated. Culture #2 is anoxic. After 16 hours the following observations are made: ■ Culture #1 has a high cell density; the cells appear to be in stationary phase, and the glucose level in the medium is reduced to 1.2%. ■ Culture #2 has a low cell density; the cells appear to be in logarithmic phase, although their doubling time is prolonged (over 1 hour). The glucose level is reduced to 0.2%. Why does culture #2 have so little glucose remaining relative to culture #1, even though culture #2 displayed slower growth and has less biomass?You are cultivating Escherichia coli in a chemostat culture. The maximum specific growth rate (µmax) of E. coli that you can reach is known as 1.0 h-1 at your culture conditions. Describe what you would observe for each condition if you have the following settings: F (flow rate of the fresh medium into the bioreactor vessel) (L/h) V (Volume of the liquid culture in the bioreactor vessel) (L) a) 1 1 b) 5 4 c) 1 2Nutrient Agar (NA) is a general purpose medium used for the cultivation of a wide variety of non- fastidious microorganisms (Merck, 2000). Its ingredients are listed in Table 2. You are tasked to prepare 300ml of NA, determine the amount of ingredients and write in column 3 in Table 2. Show your calculations. *A sample calculation was made for you. Peptone = 300 ml x 0.5% = 1.5 g Table 2. Medium Composition of Nutrient Agar (NA) with the recommended proportions (Merck, 2019) Ingredients In % In Grams/300ml Peptone 0.5 *1.5 Meat extract 0.3 Agar 1.5 Distilled Water As needed
- If the volume of a Staphylococcus aureus cell is estimated at 0.5 μm3, how many cells could be accommodated, in principle, in 5 mL of saturated culture? (1 mL = 1 cm3). Show your calculations.When the yeast cells have completely re-hydrated, measure out 1.8 mL of well-mixed yeast suspension (0.2% yeast) into each of two new 15 mL centrifuge tubes. Add 200 μL of a 10% (w/v) Sodium Azide*** in YGM solution to one of the 15 mL tubes with yeast suspension. Add 200 μl of YGM to the other 15 mL tube of yeast suspension for your Control. Let both tubes incubate at room temperature for 30 min, vortexing every ~5 minutes What is the % concentration of azide in this sample during metabolic inhibition?What is the % concentration of yeast in this sample during metabolic inhibition?Nicotiana tabacum cells are cultured to produce a polysaccharide gum. The reactor used is a stirred-tank reactor with an initial volume of 100 L. The maximum specific rate of growth of the culture is 0.18 d-1 and the yield coefficient of substrate in biomass is 0.5 gX/gS. The concentration of the limiting substrate in the medium is 3% (m/v). The reactor is inoculated with 1.5 g/L of cells and operated in batch until the substrate is exhausted, when the medium is fed with a constant flow rate of 8 L/d. The fed batch occurs in a quasi-steady state condition.a) Estimate the time of the batch step and the concentration of cells reached in this phase, considering exponential cell growth.b) The fed batch phase is carried out for a period of 40 days. What is the final concentration of cells in the reactor?c) The bioreactor is available for the process for 275 days a year, with an interval of 24 hours between each cultivation. What is the most advantageous operating mode for the process…
- Prior to subculture, Tifa used a hemocytometer to count her HepG2 cells cultured on a T-25 flask. After trypsinization, she prepared her cells by mixing 50 µL of the cell suspension with 50 μL of 0.4% trypan blue. Her observation under the microscope is as shown below: 1 (1) (ii) 3 (iii) 2 (iv) 4 Note: Count cells on the four labelled squares. Include cells touching the line on top and left. 18P 06. ¿66 Ⓡ Determine the number of viable and dead cells from her observation. What is the percentage of viability of this culture? Show your calculations in detail. Calculate the concentration of viable cells per mL in the original culture. Show your calculations in detail. Based on your understanding of cell culture, do you think she should proceed with subculture? Justify your answer.(b) A Food material containing Bacillus stearothermophilus PS1518 as an indicator organism ts subjected to heat sterilization at 121 C. Calculate the time required to reduce the organism to one tenth of the original number. Fo value for the organism is 4 minutes and the decimal reduction time , D, at 116°C is 40 minutes. Assume operation is at constant temperature of 121°C. HINT Fo = -To 10 dt Where Fo = equivalent exposure time at 121°C of the actual exposure time at a variable temperature To = the reference temperature =121 C, z=10 = number ofC necessary for10fold increase in FExplain how to prepare 250 ml liquid and 500 ml solid form separately from LB (Luria Bertani) medium whose components and concentrations are given below. Yeast extract 5 g/L Peptone 10 g/L NaCl 10 g/L Agar 1.5 g/L