Mutants Simon Bishop 613545 An organism or cell with a mutation. A mutation is a DNA base pair change or chromosome change. Mutants defective in a biological process can be used to increase our understanding of that process.
Point Mutations Missense mutation Nonsense mutation Neutral mutation Silent mutation Frameshift mutation Fig 19.3 from Essential iGenetics, Russell Mutants can be studied by selecting for known phenotype or genotype .
Share homology with Humans
Easier to study as fewer ethical regulations and shorter life cycle
BUT hard to study development
Single celled eukaryotes
Fewer control mechanisms?
Share cell division mechanism
1. The Cell Cycle Engine
Temperature-sensitive mutants of Schizosaccharomyces pombe selected which initiate M-phase at a reduced size to wild type
Led to studies of wee and cdc phenotypes
Led to discovery of mechanism for M & S phase regulation.
Has practical uses in study of Cancer.
Paul Nurse is former Chief Executive of Cancer Research UK.
Figure 2 from Nature vol 256 (1975) p549
2. Knockout organisms i
Gene Replacement: mutant allele replaces wild type, avoiding the so-called position effect
Gene Knockout: specific transgene replaces homologous gene in genome, replacing or removing gene function.
2. Knockout organisms ii Exon Neomycin R Intron Knockout chromosome (vector DNA) Chromosome of ES cell Plate on medium with Neomycin wt
2. Knockout Organisms iii Diagram from Introduction to Genetic Analysis, Gelbart, W et al Parents F1 F2
2. Knockout Organisms iv
Mlh3 belongs to a family of proteins known to have roles in meiosis and DNA mismatch repair, but the function of Mlh3 itself was unknown.
Mendelian ratios of F2 mice were as expected so Mlh3 not required for development.
F2 generation sterile .
Figure 1a from Nature Genetics vol 31 p385
2. Knockout Organisms v
Male testes were smaller with no viable spermatocytes
Female ovaries unaltered but no viable oocytes
Mlh3 shown to be required to hold chromosomes together at chiasmata during meiosis.
Mlh3 shown to be essential for reproduction and meiosis.
Figure 4g,h from Nature Genetics vol 31 p385
Two methods of mutant study:
Selection by phenotype ( wee1 mutants)
Nurse, P. Nature vol 256 (1975)
Selection by genotype ( Mlh3 ), using knockout methods