BSC 310 Lecture Notes - Lecture 21: Beta Sheet, Dimethyl Sulfoxide, Environmental Factor

13 views7 pages

graysardine594

22 Mar 2016

School

University of Alabama

Department

Biological Sciences

Course

BSC 310

Professor

Julie Olson

For unlimited access to Class Notes, a Class+ subscription is required.

Document Summary

Viable cell- one that is able to divide and form offspring. Viable counting methods rely on this ability to grow. Plate count- viable cell count on an agar medium; assumes that each single cell divided and grew to yield a single colony (cid:1) Spread-plate method- a volume of diluted culture is spread over the surface of an agar plate and incubated. Pour-plate method- a volume of culture is placed into an empty. Petri plate, molten agar is added and the culture and agar are mixed well by swirling (cid:1) (cid:1) (cid:1) Need to have a countable number of colonies, not too many and not too few (cid:1) (cid:1) Preferred range is between 30 and 300 colonies. Most samples will need to be diluted in order to return colonies within this range but because the concentration is often unknown, multiple dilutions are used (cid:1)

Related Questions

Viable cell counts are often performed on exponentially growing cells in order to relate the absorbance reading (AU) with actual cell number. For this purpose, a sample of the culture is removed during the exponential phase after the absorbance reading is recorded. Spread plates are prepared in order to perform a viable cell count. The goal is to have a spread plate containing between 30 and 300 cells (colonies) when plating a volume of 0.1 ml (100 Î¼l). To determine the correct dilution, you will need to estimate the number of cells/ml present from the AU (absorbance units) value of your sample. For this calculation, we will assume that 1 AU is equivalent to 10⁷ cells /ml. So:

AU of sample x [(10⁷ cells/ml)/1 AU] x 0.1 ml plated = total # of cells expected

QUESTION 1:

You have removed a volume of a culture of E. coli during exponential growth in order to measure the viable cell count. If the absorbance reading was 0.3, estimate the maximum dilution factor you would need in order to obtain between 30 and 300 colonies on a plate spread with 0.1 ml of the dilution. Assume that there are 10⁷ cells/ml in one absorbance unit.

QUESTION 2:

You have removed a volume of a culture of E. coli during exponential growth in order to measure the viable cell count. If the absorbance reading was 0.3, estimate the minimum dilution factor you would need in order to obtain between 30 and 300 colonies on a plate spread with 0.1 ml of the dilution. Assume that there are 10⁷ cells/ml in one absorbance unit. â

bluealligator624

In a quadrant streak, why do you intersect the previous lines and not the ends of the streak?

Select one:

a. More cell amount variability at ends, usually higher cell population

b. Keep streaks in the middle of the plate

c. Prevent cell death

d. To maintain a mixed culture

e. To count CFUs

How should you hold the inoculating loop?

Select one:

a. Maintain it at an angle at all times

b. In a test tube rack at all times

c. Like a pencil

d. Balanced on top of the tube

e. With dish gloves

Which of the following is used in the spread plate and not the streak plate method?

Select one:

a. Glass spreading rod

b. Flame loop

c. Agar plate

d. Bunsen burner/sterile utensil

e. Starting culture to sample

What would likely happen if the inoculum was not spread adequately over the agar surface?

Select one:

a. Contamination

b. Degradation

c. Sterilization

d. Uneven distribution

e. Cells will not grow

You have a mixed culture of unknown bacteria and want to identify one of the species by several subsequent experiments (needing multiple colonies of a pure culture). You do a quadrant streak and transfer to a sterile liquid medium. What next step makes the most sense?

Select one:

a. Perform a Gram stain

b. Note all colony morphology characteristics

c. Use the spread plate method or new streak plate on an isolated colony of the plate

d. Quantify the cells

e. Repeat using the other two methods of isolation

What would likely occur if the flame loop was not sterilized between streaks on the quadrant streak method?

Select one:

a. Bacteria will not stick to the loop

b. Virus will be introduced

c. Flame loop will be too hot

d. The sample will be contaminated

e. Subsequent streaks would have more colonies

Why is the glass rod in the spread plate method put in alcohol and ignited instead of sterilized like a flame loop?

Select one:

a. Glass oxidizes

b. Glass melts

c. Glass takes longer to cool

d. Glass burns

e. Glass is more sterile

Determining Dilution Scheme

Viable cell counts are often performed on exponentially growing cells in order to relate the absorbance reading (AU) with actual cell number. For this purpose, a sample of the culture is removed during exponential phase after the absorbance reading is recorded. Spread plates are prepared in order to perform a viable cell count. The goal is to have a spread plate containing between 30 and 300 cells (colonies) when plating a volume of 0.1 ml (100 ?l). To determine the correct dilution, you will need to estimate the number of cells/ml present from the AU (absorbance units) value of your sample. For this calculation, we will assume that 1 AU is equivalent to 10⁷ cells /ml. So:

AU of sample x [(10⁷ cells/ml)/1 AU] x 0.1 ml plated = total # of cells expected

You want to calculate two different dilutions that will give you between 30 and 300 colonies (cells). In order to determine the maximum dilution factor, divide the total # of expected cells by 30. In order to determine the minimum dilution factor, divide the total # of expected cells by 300. For example if the AU was 0.3, the total number of expected cells would be 3.0 x 10⁵. In this case the maximum dilution factor would be 10⁴ (3.0 x 10⁵/30) or a dilution of 10^-4 and should produce approximately 30 colonies. The minimum dilution factor would be 10³ (3.0 x 10⁵/300) or a dilution of 10^-3, which should produce ~300 colonies. Since performing a single dilution of 10^-4 or 10^-3 would be highly inaccurate, serial dilutions are performed. Typically, the spread plates are prepared in duplicate for greater accuracy.

(NOTE: Do not put units in your answers, if you do the computer will grade the answer as wrong and you will lose the points. Answers should be written in numerical form or if you want to write as scientific notation you can write as 1.22E8 or 1.22E-8. The system will only take up to 15 significant figures.) (Another NOTE: Your answer must be within 5% of the correct value to get credit for this problem.)

You have removed a volume of a culture of E. coli during exponential growth in order to measure the viable cell count. If the absorbance reading was 0.8, estimate the maximum dilution factor you would need in order to obtain between 30 and 300 colonies on a plate spread with 0.1 ml of the dilution. Assume that there are 10⁷ cells/ml in one absorbance unit. Answer:

You have removed a volume of a culture of E. coli during exponential growth in order to measure the viable cell count. If the absorbance reading was 0.8, estimate the minimum dilution factor you would need in order to obtain between 30 and 300 colonies on a plate spread with 0.1 ml of the dilution. Assume that there are 10⁷ cells/ml in one absorbance unit. Answer:

coffeecricket768

BSC 310 Lecture Notes - Lecture 21: Beta Sheet, Dimethyl Sulfoxide, Environmental Factor

Document Summary

Get access

Related Documents

BSC 310 Lecture Notes - Lecture 77: Absorbance, Exponential Growth, Archaea

BIOL240 Lecture Notes - Lecture 19: Petri Dish, Microbiological Culture, Turbidity

300833 Lecture Notes - Lecture 8: Serial Dilution, Logarithmic Scale, Turbidity

Related Questions