Lab 2 – Preparation of Nutrient Agar Plates and Aseptic Transfer of Cultures

Ninoska Garcia-Ortiz 063 053 2

Lab date: Wed, Aug 28, 2007

Objective:

To learn how nutrient agar is prepared and how to pour plates aseptically, how to transfer bacteria
using aseptic techniques; to become aware of the presence of microorganisms in the environment.

Results:

SAMPLE EXPECTED RESULTS OBTAINED RESULTS

Aseptic transfer with an No growth is expected as we

autoclaved culture of E.coli on were informed that the E.coli No Growth
to sterile Petri plate. sample was “dead”

Inoculated sterile Petri plate Growth is expected as our Growth exhibited

with finger tips fingertips are non sterile

Discussion Questions:

1. Agar – base derived from seaweed; a complex polysaccharide derived from a marine alga
(Gelidium) and used as a solidifying agent in culture media.

Bacterial Colony - a cluster of organisms growing on the surface of or within a solid

medium, usually cultured from a single cell. Bacterial growth is the reproduction of
bacteria through binary fission, in which a single cell divides in to two identical cells.

Autoclave – chamber that brings steam temperature higher than 120 degrees Celsius. It’s
a pressurized device designed to heat aqueous solutions above their boiling point to
achieve sterilization.

2. Agar by itself is not a nutrient, it is only after adding such things as beef extract and
peptone (form of protein) that one obtains nutrient agar.
 3. They are incubated upside down so that if there is any condensation that forms on the
cover it will not drip on to the cultures which would allow the organisms to spread
inhibiting the growth of colonies.

37˚C is the optimum growth climate condition for mesophiles.

4. Do not to touch rim of glass for risk of contamination when pouring

• Do not pour close to a draft because air currents could blow in microorganisms
• Do not to leave the lid open too long for bacteria can get in via air particles
• When lifting the lid to pour keep in relatively close to the plate because the further
away you take it the greater the risk for contamination.
• When pouring bring the Petri dish close to the end of the table so that you can pour
close to the dish itself which in turn minimizes the risk of splashing.

5. The sterilization procedure for autoclaving is 15min. in the autoclave and 30 min cool
down time; so about 45 min in total.
Temperature: 120 degrees Celsius
Pressure: 15 lbs/in2
Duration: 15 minutes

6. A prion, an infectious agent composed only of protein, which affect the structure of the
brain or other neural tissue, is sterilized by the denaturation of the protein to a state where
the molecule is no longer able to induce the abnormal folding of normal proteins. This is
accomplished by subjecting them to a temperature of 134 degrees Celsius for 18 minutes
in a pressurised steam autoclave.

Conclussion

In conclusion, the objectives of this experiment were met. Through careful manipulation two
aseptic plates were successfully poured. This was proven by the absence of growth on these
plates. Aseptic technique was also used during the transferring of E.coli bacteria. There was
absence of growth in this third plate as well, which was expected, as the E.coli bacteria in the
broth was dead. The results for the fourth Petri dish occurred as expected. My fingers were dirty,
not aseptic, and the micro-organisms colonized on the plate proved what I expected