Download - Modeling Transformation
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Modeling Modeling TransformatTransformat
ionionWhat does each What does each
step do?step do?
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Transformation Transformation ProcedureProcedure
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Transformation Transformation ProcedureProcedure
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Building Your ModelBuilding Your Model
Yarn Yarn = chromosomal DNA= chromosomal DNABeads Beads = ribosomes= ribosomesBlack Black velcro = plasmid DNAvelcro = plasmid DNAGreen Green velcro = GFP genevelcro = GFP genePink Pink velcro = Ampicillin resistance velcro = Ampicillin resistance
genegeneBottle = cell membraneBottle = cell membranePipe cleaner = flagellumPipe cleaner = flagellumCup = micro-tubeCup = micro-tube
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pGLO PlasmidpGLO Plasmid
GFPGFP: : gene for gene for Green Fluorescent Green Fluorescent ProteinProtein
araCaraC: : gene for gene for Beta-lactamase Beta-lactamase (Ampicillin (Ampicillin resistance)resistance)
Real image of a bacterial plasmid (Scanning electron micrograph)
GFP
araC
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E. coli E. coli CellCell
Bacterial chromosomal DNA
Cell membrane
-
-
---
-
-
- - - -
- - -Ribosomes
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Add transformation Add transformation solutionsolution
Tube
CaCl2
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Transformation Transformation solution: CaClsolution: CaCl22
Positive charge of CaPositive charge of Ca++++ ions shields negative ions shields negative charge of DNA charge of DNA
OCH2
O
P O
O
OBase
CH2
O
P
O
O
O
Base
OH
Sugar
Sugar
O
Ca++
Ca++
Ca++
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Inoculate with bacteriaInoculate with bacteria
Tube
CaCl2
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Add plasmidAdd plasmid
Tube
CaCl2
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Incubate on iceIncubate on ice
Slows movement of cell Slows movement of cell membranemembrane
pGLO plasmids
Bacterial chromosomal DNA
Cell membrane
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Heat-shock @ 42°CHeat-shock @ 42°C
Increases Increases permeabilitypermeability of of membranemembrane
pGLO plasmids
Bacterial chromosomal DNA
Cell membrane
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Incubate on iceIncubate on ice
Returns bacteria to temperature Returns bacteria to temperature necessary for survivalnecessary for survival
pGLO plasmids
Bacterial chromosomal DNA
Cell membrane
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Add nutrient broth Add nutrient broth (LB)(LB)
Provides nutrition for growth Provides nutrition for growth and gene expressionand gene expression
pGLO plasmids
Bacterial chromosomal DNA
GFP
Beta lactamase(ampicillin resistance)
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What do you expect to What do you expect to happen?happen?
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What do you expect to What do you expect to happen?happen?
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What do you expect to What do you expect to happen?happen?
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What do you expect to What do you expect to happen?happen?
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What do you expect to What do you expect to happen?happen?
-pGLO
+pGLO
LB LB/amp
LB/amp
LB/amp/
arabinose