1. Forward Genetics What is forward genetics?
Genetic screens designed to find genes that affect a trait of interest.
PHENOTYPE GENE
Random mutagenesis
Screen for interesting phenotypes
Track down the gene(s) responsible
Traits can include morphology, physiology, behavior
Reverse genetics?
Genetic screens designed to identify phenotypes association with disruption of a particular gene or DNA sequence of interest.
GENE PHENOTYPE

2. Forward Genetics Retinotectal projections in zebrafish Wild type
never mind
belladonna
astray
Baier et al., Kalstrom et al., and Trowe et al., 1996, Development

3. Saturation mutagenesis
Large scale genetic screens that aim to create a mutation in every gene required for a trait.
How do you know when you get there? When new mutations represent a second or multiple hit in a previously identified gene.
The saturation point defines the set of genes required INDIVIDUALLY for a particular trait.
Genes whose function are redundantly provided by another loci will be missed.

4. Mutagens Radiation Chemicals Insertions + Strong mutations –
deletions, inversions,
translocations
+/- May disrupt multiple
genes
Can be difficult to clone
+ Full spectrum of
mutations, usually small
+ Randomly distributed
- Difficult to clone
+ Full spectrum of mutations
+ Mutation is tagged, and easy to clone
+ Often reversible
- Nonrandom distribution

5. Genetic Screens Screen for dominant mutations (F1 screen)
Screen for recessive mutations (F3 screen)
* ¼ of families will produce m/m offspring!

6. Genetic Screens

7. ? ? Classification of Mutants: Complementation
If you have two mutants that have the same phenotype, how to test if they are mutations in the same or different genes? ? ?
Complement: provide the function of
Complementation group: a set of alleles that fail to complement one another
Complementation testing reduces a large number of mutations to a smaller set of loci.
Non-allelic non-complementation can occur if mutations affect genes in the same pathway
Hartl and Jones, 1998

8. Classification of Mutants: ComplementationX X
Mutant phenotype
Non-Complementation
Mutations are in the same gene
WT phenotype
Complementation
Mutations are in different genes

9. Classification of Mutants: The ‘Morphs’
Loss of Function alleles
Amorph
null allele, no WT activity.
genetic definition m/m = m/Df
Hypomorph
partial l-o-f, reduced WT activity.
Usually recessive, however can be dominant if it is haploinsufficient (eg mutations on the X in males)
genetic definition m/m < m/Df

10. Classification of Mutants: The ‘Morphs’
Gain of function alleles
Hypermorph
extra WT function (usually dominant)
Examples include constitutive promoters, constitutively active proteins
genetic definition: m/m> m/+ > m/Df
Neomorph
novel function (usually dominant)
Examples include chimeric proteins due to translocations
genetic definition: additional alleles (+ or Df) don’t affect the phenotype. So m/+ = m/Df

11. Classification of Mutants: The ‘Morphs’
Gain of function alleles
Antimorph
dominant negative
protein product antagonizes (poisons) the WT protein.
Occurs with proteins that function as multimers.
B-galactosidase functions as a tetramer
Wild Type
Mutant

12. Molecular Characterization of Mutants
Chromosomal Mapping (recombination mapping)
Molecular identification
Positional cloning
Rescue with candidate genes
Identification of altered transcriptional profiles
Cloning based for insertions