Closed Growth Curve Lab Report

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Microbiology (Lab Report)
Closed system growth curve

Closed System Growth Curve
Lab Report
PURPOSE
Bacteria grown in a closed system show a specific growth pattern called the growth curve which consists of four phases. The lag phase, which is a period of slow growth; exponential phase, period of maximum growth; stationary phase, where nutrients become the limiting factor making the growth rate equal to the death rate and the death phase where organisms die faster than they are replaced. It is important to know how fast a microbe grows in order to know bactericidal or antibiotic concentrations, temperature and ph at which they stop growing. Evaluating a growth curve gives you a perspective of the generation time and the mean growth rate constant which would help you estimate the minimum, maximum and optimum growth temperature of the microbe. In this experiment you would be able to see the duration of each phase, the mean growth rate constant (k) is used to measure how fast cells are dividing in a culture, generation time, optical density (OD) which s the measure of the amount of light absorbed by a suspension of bacterial cells, and the organism’s minimum, maximum, and optimum temperatures MATERIALS

* Five hot water baths, each set at different temperatures as followed; 22⁰C, 25⁰C, 31⁰C, 37⁰C and 40⁰C * Five thermometers to monitor and record temperatures in the baths * 5 side arm flasks with 47 mL of sterile Brain Heart Infusion broth + 2% NaCl * 5 side arm flasks with 50mL of sterile Brain Heart Infusion broth + 2% NaCl * Lab tissue( kim whipes)

* Micropipettes with sterile tips
* Alcohol flame or Bunsen burner
* 20mL overnight broth cultures of E.Coli in sterile Brain Heart Infusion broth + 2% NaCl PROCEDURE
1. Gather your spectrometer, micropipettes and tips, side-arm flasks with 47mL of sterile Brain Heart Infusion broth + 2% NaCl as well as the 50mL. 2. Label all the side-arm flasks containing 50mL “control” and the side-arms containing 47 mL “growth flask” 3. Put the five “growth flasks” into separate water baths for 15 minutes. 4. Turn the spectrometer on and let it warm for a few minutes. Make sure you set to “absorbance “and the wavelength to 650 nm. 5. After the flasks are mix the E.coli and add 3 drops of the mixture into separate growth flasks. Clean the and wipe the side arm and quickly take the turbidity reading. 6. Then record the observed absorbance as well as the temperature and place the flask back into its corresponding bath. 7. Monitor temp carefully. Record turbidity reading, absorbance every 15 minutes. Be consistent in tempt and position when recording absorbance.

V.RESULTS

LAG PHASE
EXPONENTIAL PHASE

CALCULATIONS
K= log (3) -- log (4.45 x 10^8) = -8.17 -8.17/ (0.301 x 12) =-2.26 k=-2.26 g =1/ (-2.26)= -0.44 g=-0.44l

Lag phase
Exponential Phase
Stationary phase
Death phase

CALCULATION
K=log(4.4)-log(0.87)=0.700.70/(00.301x8)=0.29 k=0.29
g=1/0.29=3.45g=3.45

Table 1: OD600 reading and Cell Densities
Time (h):| Viable cells/ml:| O.D. 600:|
0.5| 3.1x101| 0.63|
1| 3.2x101| 0.65|
1.5| 8.4x101| 0.87|
2| 2.58x102| 1.08|
2.5| 8.57x102| 1.37|
3| 2.11x103| 1.55|
3.5| 9.80x103| 1.9|
4| 2.04x104| 2.04|
4.5| 5.92x104| 2.28|
5| 1.90x105| 2.53|
5.5| 3.15x105| 2.62|
6| 1.71x106| 3.03|
6.5| 4.33x106| 3.21|
7| 1.42x107| 3.45|
7.5| 3.11x107| 3.64|
8| 8.70x107| 3.76|
8.5| 3.39x108| 4.16|
9| 1.04x109| 4.4|
9.5| 1.04x109| 4.38|
10| 9.68x108| 4.4|
10.5| 1.04x109| 4.4|
11| 1.03x109| 4.4|
11.5| 1.04x109| 4.4|
12| 4.45x108| 4.21|
18| 3.49x102| 1.18|
24| 3| 0.13|

| 10-1| 10-2| 10-3| 10-4| 10-5| 10-6|
6 hours 3pm| TNTC| 587| 312| 171| 105| 75|
8 hours 5pm| TNTC| TNTC| TNTC| 871| 87| 8|
11 hours 8pm| TNTC| TNTC| TNTC| 923| 301| 103|
*Note: TNTC...
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