Objective:
To find the generation time of given
organism by turbidity method.
Principle:
Turbidometric
method is the rapid, sensitive and useful technique to measure the growth of
microbes. Turbidometry is based on the fact that microbial cells scatter light
striking them. Since the microbial cells in a population are of roughly
constant size, the amount of scattering is directly proportional to the biomass
of cells and indirectly related to cell number. One visible characteristic of
growing bacterial culture is the increase in turbidity (cloudiness) of the
medium. When the concentration of
bacteria increasing, the medium appears slightly cloudy or turbid. Further
increase in concentration results in greater turbidity. When a beam of light is
passed through a turbid culture, the amount of light transmitted is measured,
Greater the turbidity, lesser would be the transmission of light through
medium. Thus, light will be transmitted in inverse proportion to the number of
bacteria. Turbidity can be measured using spectrophotometer.
Materials
required:
-Nutrient broth
- 10- 12 hrs old
broth culture
-Spectrophotometer
-Cuvette
Procedure:
- Prepare and take 50ml of nutrient broth in
sterile flask.
- Then this broth is inoculated with 1 ml of
overnight bacterial culture and places it in incubator.
- The 0th hour reading take by using
spectrophotometer and the culture is again keep it in incubator.
- The flask is removing from incubator for a
particular interval time (30 min.) and the sample is transfer aseptically to
the cuvette using sterile pipette for observe the reading.
- The optical density reading in spectrophotometer
is at 620nm.
- The reading will be tabulated and plotted the
graph by OD value in Y-axis and incubation time in X -axis.
Determination of generation time:
-Select the 2 points on the optical density scale
that represents doubling the turbidity.
The generation time is to be determine by using the
formula
Gt = t (OD2) –t(OD1)
Result:
The generation time of _______cells found
to ________ min/hr.
Peptone Broth Composition:
Peptone
20g
Sodium chloride 5g
Distilled water 1000ml
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