Table of Contents
Why Do Genetics
Genetic Terms
More Terms
Basic Molelcular
Biology
More Basic Concepts
Screens
Selections
Mutation Frequency
Chemical Mutagenesis
Frameshift Mutation
DNA Repair
Mutation Summary
Detecting Mutants
Complex Mutation
Insertion Sequences
Compound Transposons
Complex Transposons
Models of
Transposition
Transposition Summary
Mutagenesis in vitro
Effects of Mutations
Complementation
Plasmids and
Conjugation
F Factor
Transformation
Transduction
Generalized
Transduction
Specialized
Transduction
Complementation
Mapping
Two Factor Crosses
Deletion Mapping
Other Mapping Methods
Strain Construction
Inverse Genetics
Gene Isolation
Characterization of
Clones
Sequence Data
General Approaches
Fusions
Supression
Final Summary
Problem Set 1
Problem Set 2
Strain construction
©2003 written by Gary Roberts, edited by Timothy Paustian, University of Wisconins-Madison
VIII G. STRAIN CONSTRUCTION
In general, this text has treated strains with more than one mutation as if they are always a "given". In practice, of course, these need to be built. In many cases, strains can be obtained from other researchers, of course, but they had to have been "built" by someone.
The addition of a desired mutation to a strain already possessing one or more other mutations can be accomplished several ways. If the desired mutation is selectable, then it can be selected following either mutagenesis or transfer of the mutation from another strain. If it is not selectable it can be found:
By screen, following mutagenesis.
By screen, following gene transfer from another strain (where the recipient carries a linked, deleterious mutation, the cross selects for the wild-type allele of that gene from the donor and an appropriate fraction of the recipients receive the desired mutation)
By selection/screen following gene transfer from the donor that already contains a selectable insertion within the gene of interest; this latter marker (typically a transposon) is selected, but because of transposition events, not all selected strains will carry the desired insertion. A screen for the desired class completes the construction.
By selection/screen following gene transfer from a donor containing the desired mutation with a linked selectable marker like a transposon.
The above description should make clear the power of transposons in genetic analysis. Also, the introduction of a desired mutation is only part of the game--there are certainly mutations in the recipient that you do not wish to lose as well as donor markers that you do not wish to inherit. For these reasons, the "gene transfer" referred to above should be by a system which only moves relatively small portions of the chromosome; transformation and generalized transduction tend to be the systems of choice.
