Griffith’s Mysterious Transformation Experiment

Transformation

• DNA is released into surrounding medium• Recipient cells incorporate it into themselves

from the medium

How does it work?

*plasmids*

Bacterial chromosomal DNA

Cell membrane

E. Coli = Escherichia coli • Normally harmless bacteria in your gut• E. Coli can be artificially transformed by exposure to calcium chloride solution and

thermally “shocked” to become receptive to foreign plasmid

What are we doing?• We will transform bacteria (E. coli),

giving it the ability to make green fluorescent proteins as well, make it ampicillin resistant.

Making Plasmids

Gene

Plasmid

+

=

pGLO Plasmid

GFP

araC

To make green fluorescent proteins

To survive with Ampicillin (antibiotic)

• Bacteria Colonies

• Lawn – all together not separate

4 plates• Luria Broth no plasmid (-plasmid

LB)• Luria Broth With plasmid (+plasmid LB)

• AMP Broth No plasmid (-plasmid LB/AMP)

• AMP Broth With plasmid (+plasmid LB/AMP)

What do you expect• Luria Broth no plasmid (-plasmid LB)Normal growth = lawn of bacteria

• Luria Broth With plasmid (+plasmid LB)Normal growth = lawn of bacteria• AMP Broth No plasmid (-plasmid LB/AMP)NO growth, all killed• AMP Broth With plasmid (+plasmid LB/AMP)Individual Colony growth of transformed

resistant bacteria

Results

http://www.phschool.com/science/biology_place/labbench/lab6/concepts1.html

• Sterile technique!!• Don’t touch anything• Lift Petri lid partially, pour/swab etc.

Immediately reclose• Open instruments and use

immediately• Do NOT touch working end• Close quickly• Incubate ndsᴉpǝ poʍu!!