Culturing Microorganisms RPA

Question 1

What is the aim of the investigation

Answer 1

Investigate the role of antibiotics on bacterial growth using an Agar plate
(And how to prepare an uncontaminated bacteria culture first using the aseptic technique)

Question 2

Method for RPA

Answer 2

1) Spray work surface with disinfectant and dry with paper towel
2) use a pencil to separate the bottom of the Agar plate into 3 segments and put a dot in the middle of each one
3) Wash hands
4) flame loop then use loop to spread bacteria over Agar plate
5) Use forceps to place each different sterile filter paper disc containing the antibiotic on each dot
6) Tape lid on the Agar plate then incubate at 25 degrees for 48 hours
7) measure area of each inhibition zone using a ruler and the formula pie r squared

Question 3

What aseptic techniques are used to avoid contamination

Answer 3

Flame loop (before using it to spread bacteria onto Agar plate)
Use disinfectant to spray surfaces clean
Wash hands
Only open the lid of the dish when necessary and lift at an angle
(And open lid by Bunsen burner flame to kill any bacteria in the air)

Question 4

What are the areas called where the bacteria hasn’t grown

Answer 4

Zone of inhibition

Question 5

Why only loosely tape lid onto Agar plate before incubating

Answer 5

To allow oxygen to reach bacteria
(Which prevents the growth of harmful anaerobic bacteria)

Question 6

What temperature should the Agar plate be incubated at and for how long

Answer 6

25 degrees
48 hours

Question 7

Why is the Agar plate incubated at 25 degrees

Answer 7

Prevents growth of harmful microorganisms

Question 8

How to accurately measure area of inhibition zone

Answer 8

Use ruler to measure diameter then measure again at 90 degrees and calculate a mean diameter
Divide by 2 to get radius then do pie r squared

Question 9

2 sources of error

Answer 9

Contamination from other bacteria may have occurred
Irregular shaped inhibition zone is difficult to determine width from