Mutations include all those heritable changes, which alter the phenotype of an individual. Hugo de Vries used the term mutation to describe the phenotypic changes which were heritable.
Mutation was first discovered by Seth Wright in 1791 in male lamb having unusual short legs. The scientific study of mutations started in 1910, when T.H. Morgan started his work on fruitfly, i.e., Drosophila melanogaster.
Types of Mutations
A classification based on the method of detection of mutations includes the following main types:
- Morphological mutations involve alterations in external form including colour,shape, size etc. Examples include albino ascospores in Neurospora, kernel colour in corn, curly wings in Drosophila and dwarfism in pea.
- Lethal mutations involve genotypic changes leading to death of an individual.
- Biochemical mutations are identified by a deficiency, so that the defect can be overcome by supplying the nutrient or any other chemical compound, for which the mutant is deficient. Such mutations have been studied mainly in prokaryotes such as bacteria and fungi, but sometimes also in eukaryotes such as Drosophila and humans.
- Resistant mutations are identified by their ability to grow in the presence of an antibiotic (e.g., streptomycin, ampicillin, cycloheximide) or a pathogen, to which wild type is susceptible.
- Conditional mutations are those which allow the mutant phenotype (including lethality) to express only under certain condition (e.g., high temperature) termed as restrictive condition.
Mutation Rates and Frequencies
Mutation rates at individual loci
The mutation rates, per gene replication or per cell division could be measured in organisms such as bacteriophages, bacteria or unicellular algae such as Chlamydomonas, where from the number of mutant cells and normal cells, an estimation of number of cell divisions can be made.
Mutation frequencies at individual loci
The frequencies are more conveniently estimated. Different genes in an organism differ in mutation frequencies. Some genes are unstable or mutable and thus mutate more frequently than others. L.J Stadler used dominant marker stocks as male parents and crossed them with female recessive stocks to estimate mutation frequencies. Similar estimates of mutation frequencies were also made in fruit fly (i.e., Drosophila), mouse and human.
Mutation frequency per genome
Mutation frequencies can also be expressed in terms of whole genome irrespective of genes involved. For instance, in Drosophila, an estimated number of genes is 5,000 and since average mutation frequency at a particular locus is one in 100,000, we should expect that one out of every 20 flies should have a mutation. In man this frequency is perhaps higher.
Physical and chemical mutagens are the main mutagens for inducing mutations artificially.
These are mainly radiations, although change in pH value (acidity) or temperature shocks may also induce mutations.
Ionizing radiations will cause ionization and will force ejection of an electron from the atom it attacks. But the non ionizing radiations such as UV do not cause ionization, but cause excitation through energy transfer. Among ionizing radiations more commonly X-rays, gamma rays, beta rays and neutrons are helpful for inducing mutations.
The chemicals used for inducing mutations were mustard gas, ethyl urethan, phenol, formaldehyde, etc. Mustard gas is highly mutagenic, having a delayed effect. Ethyl methane sulphonate (EMS) induces mutations in microorganisms, higher plants and animals.
Detection of Mutations in Drosophila
Mutations induced due to radiations or due to chemicals could be broadly of two types, lethal mutation and visible mutation. Both these types could be located either on sex chromosomes or on autosomes (autosomes are chromosomes other than sex chromosomes).
Detection of sex linked lethals
The CIB method:
The CIB method involves use of a CIB stock which carries an inversion in heterozygous state to work as crossover suppressor (C), a recessive lethal (l) on X chromosome in heterozygous state and a dominant marker, Barred (B) for the barred eye (narrow eye).
One of the two X chromosomes in a female fly carried all these three features and the other X chromosome was normal. Male flies irradiated for induction of mutations where crossed to CIB females. Male progeny receiving CIB X-chromosome will die. The CIB female flies obtained in progeny can be detected by barred phenotype. These are crossed to normal males. In the next generation 50% of males receiving CIB X- chromosome will die. The Other 50% males will receive X chromosome, which may or may not carry the induced mutation. In case lethal mutation was induced no males will be observed. On the other hand, if no lethal mutation was induced 50% males will survive. Thus, the CIB method was the most efficient method for detecting sex-linked lethal mutations.
Muller 5 method:
This method uses Muller 5 Drosophila stock, which carries two marker genes, dominant Bar ( Barred eye) and recessive apricot, but does not carry a lethal as in CIB. Moreover Muller 5 contains a more complex inversion and therefore, has a better crossover suppressor than in CIB . In F2 generation, 50% males are Muller 5 phenotype and remaining 50% are wild-type. Induction of a lethal mutation in X chromosome of irradiated male will show no wild type males in F2 generation. Therefore, absence of wild type males in F2 is an indication of induced lethal mutation.
Using different doses of irradiation and working out the corresponding frequencies of lethal mutations, we observe a linear relationship between the radiation dose and mutation frequency. Use of different kinds of radiations shows that the alpha particles gave the lowest mutation frequencies and beta, gamma and hard X rays give the highest frequency.
Detection of sex during the visible mutations:
Attached X method:
For detection of sex linked visible mutations this method requires Muller 5 and attached X chromosomes . The attached X females have a special advantage. When these females are crossed to an irradiated male, X chromosome of irradiated male goes either to super female daughters or to the sons. Since in sons there is a single X chromosome, any visible induced mutation will immediately express itself.
Detection of mutation in plants
Mutations at specific loci
Stadler’s method :
L.J. Stadler studied frequencies of spontaneous mutations in maize for endosperm characters. Following were the steps:
- Growing and detasseling of a genetic stock dominant for several genes as female parent .
- Sowing of multiple recessive stock on every 5th row to supply pollen.
- Examine the seed set on female plants for endosperm characters, example shrunken. Most of the seeds showed dominant phenotype; the number of seeds showing recessive character represented mutation in female gametes.
Stadler also induced mutations artificially. For this purpose, he used irradiated pollen from dominant stock for pollinating recessive stock. Progeny showing recessive phenotype were classified as mutants.
W.R. Singleton also studied induced mutations. The pollen from plants dominant for several genes and growing in a field with radiation source were helpful to pollinate recessive stock growing in a field without radiation source. The pollen carrying mutation will give seeds which will show recessive character in phenotype.
Mutations at unspecified loci
When we use a plant species, where no marker stocks are available and the purpose is to study visible mutations in genome as a whole, then the mutations are studied by segregation in M2 generation. It involves the following steps:
- Irradiate the seeds.
- Grow plants in M1 generation from irradiated seeds and self them.
- Harvest the seed in M1 on single plant basis.
- Grow M2 on single plant progeny basis and study segregation in M2 families. Each segregating family will represent one mutation in an irradiated seed.