Animal Models

Front 1

Transgenic mouse

Back 1

-multiple species
-insert foriegn DNA randomly into pronucleus of fertilized egg, then implant egg into foster mom
-exam w/tail biopsies-PCR or south blot
-usually-ectopic misexpression(in cells where not normally expressed)
-Or overexpression
-random insertion=variable expression

Front 2

Transgenic mouse Preperation

Back 2

need:
1)Plasmid containing-promotor(high levels or tissue specific),transgene,termination/polyA makes stable mRNA
2)fertilized egg
3)foster mom
4)other mice to mate positive pups

Front 3

Transgenic mouse applications

Back 3

A)Analysis of gene function through overexpression
B)Animal models of human disease(extension of (A),models diseases assoc. with overexpression)
C)Production of foriegn proteins

Front 4

Knock-out mice

Back 4

-mice only
-targeted insertion in ES cells
-disrupt specific genes to reveal func.
-exam-by linking hair color to gene

Front 5

Targeted insertion production

Back 5

1)Plasmid and targeting sequence must be homologous recombination-pos/neg selection
2)corectly targeted ES cells microinjected into blastocysts removed from donor female
3)put into foster pseudopregnant mom
4)mate chimeric mice to wild-type mice

Front 6

Target insertion applications

Back 6

A)Knock-out suble mutations,even point mutations to interfere with aspects of gene func.
B)Gene knock-in=replacement or alteration of endogenous gene
C)manipulate gene copy number to raise or lower level of production
D)intro of reporter gene for studies of promotor function