Transcript Lab Exercise #17

Bio 112 Genetics Lab
Exercise 17
Dihybrid Cross in Corn
There are four grain phenotypes in the above ear of corn: Purple & Starchy(A),
Purple & Sweet(B), Yellow & Starchy(C) and Yellow & Sweet(D).
These four grain phenotypes are produced by the following two pairs of heterozygous
genes (R & r and SU & su) located on two pairs of homologous chromosomes (each
gene on a separate chromosome):
Dominant alleles
Recessive alleles
R = Purple
r = Yellow
SU = Starchy
su = Shrunken
The cross which produced the corn ear above was:
Rr SUsu X Rr SUsu
DNA Extraction from Bacteria
Transformation of Bacteria
(pGLO)
We will introduce a gene into E. coli that will produce a
protein (green fluorescent protein-GFP) that will cause
the colonies to glow green when exposed to ultraviolet
light
What is Transformation?
Uptake of DNA (in this case a plasmid pGLO) from the
surrounding environment of the cell.
What is Green Fluorescent ProteinGFP)?
GFP was discovered in the bioluminescent jelly Aequorea
victoria. The gene that makes this protein is used extensively
in research…and also for fun?
Transformation Procedure … in a
nutshell.
• Suspend bacterial colonies in Transformation Solution,
CaCl2
• Add pGLO plasmid DNA to +DNA tube
• Place tubes on ice
• Heat shock at 42oC and place on ice
• Incubate with LB broth
• Streak plates
Expected Results
Our genes of interest:
amp…araC…GFP
araC
ori
pGLO
bla
GFP
 amp – this gene will give our transgenic bacteria
resistance to the antibiotic ampicillin
 araC – this gene will produce a protein which in the
presence of the sugar arabinose will allow the bacteria
to turn on the GFP gene
 GFP – in the presence of arabinose, this gene will
“turn on” and cause the transformed (transgenic)
bacteria to glow green
The Role of Arabinose
• The bacterial genes that make the digestive
enzymes needed to break down arabinose for
food are not expressed (made) when arabinose is
absent.
• When arabinose is present the genes are turned
“on”. When it is absent the genes remain “off”.
• Arabinose initiates transcription of the genes by
promoting the binding of RNA polymerase to the
promoter.
araC
ori
pGLO
bla
GFP
Expected Results