Lab 4E, 4F & 2B
Media Prep. & Model Organisms
Timeline/Overview
 Monday: Lecture
 Tuesday: Media Prep (4E & 4F)
 Pour Plates for Day 2 of Lab 2B
 Thursday: Media Prep (4E & 4F)
 Tuesday: Lab 2B—part I, II, III
 Wednesday: Lab 2B—part IV
 Thursday: Lab 2B—repeat part IV, compile data
Preparation for 4E & 4F
4E & 4F
 Lb broth base and LB agar base
 Several 250ml jars
 Bring in extra hood
 10% Bleach
Prep. For Lab 2B
Month Prior
 Check supplies: bleach, petri dishes, LB base, inoculating
loops, 3 incubators, yeast, glucose, foil
 Order: E. coli, Aspergillus, Potato plates
Week Prior
 prepare LB agar & broth during Lab 4E & 4F
 Pour plate (2 per group20)
 Set incubators at 30, 37 and 42 C
 Scalpels from other room
1 day prior
 make bacteria culture
Media Prep.
 Luria Bertani (LB) Agar.
 Use to grow bacteria and any other microorganisms.
 An all purpose agar.
 Media Prep Equation:
 Mass1/Volume1 = Mass2/Volume2
 Steps on Preparing Agar Plates
 Use a 250-mL beaker to use as a container for the agar.
 Pour LB Agar Base with Distilled water into the container and mix till the base is fully
fused with the distilled water.
 Place the container on a Heat Source (i.e. microwave) for 15 seconds and every 15
seconds stir the container until the container is fully clear from debris. Repeat this step
until the container is cleared.
 Finally, after the container is all clear from debris, place the cap of the container but, very
loosely so, it can release pressure during the autoclave. Have the cap jiggle.
 Place the container in the autoclave from 15 to 20 minutes at 15 to 20 psi.
Model Organisms
 Characteristics of Model Organisms:
 Easy to grow & maintain
 Easy to provide nutrients
 Short Generation time
 Well Understood Growth & Development
 Resembles Other Organisms
 Escherichia Coli (E. coli), Aspergillus & Yeast
 Levels of Biological Organization:
 Atoms
Molecules
Organisms
Organelles
Cells Tissues
Organs
2B purpose
To grow and study an organism in a
laboratory, one should know the
environmental preferences of the species.
Maintaining an organism at less than optimal
light intensity, temperature, or oxygen level,
from example, may put the organism under
stress, possibly affecting its growth or other
processes.
Pre-lab—Plate Pouring (4E & 4F)
 Pour LB agar base plates (Luria-Bertani )
E. Coli plate—use the LB agar with sterile technique, use a
10ml pipette to transfer 10ml into a sterile plate
Yeast plate—use the LB* agar, sterile technique, transfer 10ml
using a pippet
*we are not using Malt extract agar because we do not have it,
but LB agar should have enough nutrients to grow yeast
Each group does 1 plate of each organism at their assigned
Temperature—wrap in foil for dark
1.
2.
3.
4.
5.
6.
7.
8.
9.
room
room
30C
30C
37C
37C
42C
42C
8C (instead of the suggested 4C)—please do 2 sets
Part I Tips
 Label the bottom
 Inoculating loop
 Triple Z streaking method
 Close lids with tape
 Make sure to use plate with LB agar
Part II Tips
 Each group gets one plate instead of a tube of potato
dextrose agar
 We are using a potato dextrose agar plate NOT tube
 Cut aspergillus from stock plate—will be in the hood
 Put the aspergillus (fungi) on the plate fungi side down to the
agar and close the lid
Part III Tips
 Label the bottom
 Inoculating loop
 Triple Z streaking method
 Close lids with tape
 Make sure to use plate with LB agar
Part IV—Data Collection & Analysis
 Make sure to come in at FIRE on Wednesday to do a table




like 2.3 (add space for 8C)
We will repeat the table again on Friday in class
This will provide us with more information of what happens
over time as well as temperature
The growth is exponential, but when nutrients run out, they
go into the death phrase (smelly!), we want to try to catch
them at their peak
Use results from all the groups posted on the internet for
you data
Tutorials
 Media Prep. (13)
 LB Agar and Broth (14)
 Sterile technique (15)
 Pouring Plates (16)
 Starting a Broth (18)
 Streak Colonies (17)