Chapter 062. Principles of Human Genetics (Part 10) docx

Principles of Human Genetics Part 10 Transgenic Mice as Models of Genetic Disease Several organisms have been studied extensively as genetic models, including Mus musculus mouse, Dros

Chapter 062 Principles of

Human Genetics

(Part 10)

Transgenic Mice as Models of Genetic Disease

Several organisms have been studied extensively as genetic models,

including Mus musculus (mouse), Drosophila melanogaster (fruit fly),

Caenorhabditis elegans (nematode), Saccharomyces cerevisiae (baker's yeast),

and Escherichia coli (colonic bacterium) The ability to use these evolutionarily

distant organisms as genetic models that are relevant to human physiology reflects

a surprising conservation of genetic pathways and gene function Transgenic mouse models have been particularly valuable, because many human and mouse genes exhibit similar structure and function, and because manipulation of the mouse genome is relatively straightforward compared to those of other mammalian species

Transgenic strategies in mice can be divided into two main approaches: (1) expression of a gene by random insertion into the genome, and (2) deletion or targeted mutagenesis of a gene by homologous recombination with the native endogenous gene (knock-out, knock-in) (Fig 62-6; Table 62-3) Transgenic mice are generated by pronuclear injection of foreign DNA into fertilized mouse oocytes and subsequent transfer into the oviduct of pseudopregnant foster mothers

Figure 62-6

Transgenic mouse models Left Transgenic mice are generated by

pronuclear injection of foreign DNA into fertilized mouse oocytes and subsequent

transfer into the oviduct of pseudopregnant foster mothers Right For targeted

mutagenesis (gene knock-out/knock-in), embryonic stem (ES) cells are transfected

with the targeted (mutagenized) transgene The transgene undergoes homologous recombination with the wild-type gene After selection, positive ES cells are introduced into blastocysts and implanted into foster mothers Chimeric mice can

be identified based on the mixed coat color of the offspring Heterozygous mice are bred to obtain mice homozygous for the mutant allele

Table 62-3 Genetically Modified Animals

Commonly

Used Description

Technical Principle

Remarks

Commonly used

Genomic DNA or cDNA constructs

Random integration of transgene

Transgenic Pronuclear

injection of transgene

Variable copy numbers of transgene

Variable expression in each individual founder

Gain-of-function models due

to overexpression using tissue-specific promoters

Loss-of-function models using anti-sense and dominant negative transgenes

Inducible expression possible (Tetracycline, ecdysone)

Applicable to several species

Predominantly used in mice (Targeted)

Knock-out

Substitution of functional gene with inactive gene by homologous

Tissue-specific knock-out possible (Cre/lox)

recombination in embryonic stem cells

Absence of phenotype possible due to redundancy

Predominantly used in mice (Targeted)

Knock-in

Introduction of subtle mutation(s) into gene by substitution of endogenous gene with gene carrying a specific mutation Homologous recombination in embryonic stem cells

Can accurately model human disease

Selection of phenotype followed by genetic characterization

Forward

genetics

Mutations created

randomly by ENU (N-ethyl-N-nitrourea)

Useful for identifying novel genes

Congenic

strains

Mating of an

inbred donor strain with a

disease phenotype with an

Useful for mapping disease-causing genes

inbred recipient strain in

order to define the

responsible for the disorder

Successful in several mammalian species including sheep (Dolly), mice, cows, monkeys

Cloning of genetically identical individuals

May affect life-span

Cloning Introduction of

nucleus into enucleated eggs (nuclear transfer)

Ethical concerns