SPONTANEOUS MUTATIONS IN MICRO ORGANISMS.

SUBMITTED TO:

Dr.A.VIJAYA GOPAL

ASST.PROFESSOR

SUBMITTED BY:

B.PRASANNA KUMAR

ID.NO:RAM/14-81

Griffith's experiment, was an experiment done in 1928 by Frederick Griffith. It was one of the first experiments showing thatbacteria can get DNA through a process called transformation.[1][2]

Griffith used two strains of Pneumococcus. These bacteria infect mice. He used a type III-S (smooth) and type II-R (rough) strain. The III-S strain covers itself with a polysaccharide capsule that protects it from the host's immune system. This means that the host will die. The II-R strain does not have that protective shield around it and is killed by the host's immune system.In this experiment, bacteria from the III-S strain were killed by heat, and their remains were added to II-R strain bacteria. While neither harmed the mice on their own, the blend of the two was able to kill mice.Griffith was also able to get both live II-R and live III-S strains of pneumococcus from the blood of these dead mice. He concluded that the type II-R had been "transformed" into the lethal III-S strain by a "transforming principle" that was somehow part of the dead III-S strain bacteria.[3]

Today, we know that the "transforming principle" Griffith saw was the DNA of the III-S strain bacteria. While the bacteria had been killed, the DNA had survived the heating process and was taken up by the II-R strain bacteria. The III-S strain DNA contains the genes that form the shielding polysaccharide part from attack. Armed with this gene, the former II-R strain bacteria were now protected from the host's immune system and could kill the host.The exact nature of the transforming principle (DNA) was confirmed in the experiments done by Avery, McLeod and McCarty and by Hershey and Chase.

Definition:

A sudden and heritable change in the nucleotide sequence of a gene.

A mutation is a change in phenotype, which is sudden, heritable and is not produced due to segregation or recombination.

Mutation is the ultimate source of all the genetic variation existing in any organism.

Mutations always change the genotype and thus the phenotype of the microorganism.

Spontaneous mutations are those that arise occasionally in the absence of a known cause i.e., without exposure to external agents.

These mutations may result from errors in DNA replication, or from the action of transposons,or even from the effect of some mutagenic agents present in the environment.

In India, the first mutationbreeding programme beganin the early 1930’s.

Mutations are random events interms of the time of their occurrence and the gene inwhich they occur.

Most mutations have harmful effects, but some mutations are beneficial.

Mutations are recurrent.i.e., the same mutation may be expected to occur in different individuals of a given generation.

Mutations can occur in any tissue or cell of an organism.

Mutations occur in both forward & reverse directions. Generally the rate of forward mutations are much higher

than those for reverse mutations. Mutations occur at very low frequencies in nature such

mutations are called spontaneous mutations. The rate of spontaneous mutations for most of the genes

is very low ranging between 10⁻7 and 10⁻4. Some genes increase the spontaneous mutation rates of

some other genes of the genome. Such genes are called mutator genes.

The spontaneous mutation rate varies. Large gene provide a large target and tend to mutate more frequently.

Organisms possess a variety of cellular functions to suppress spontaneous mutagenesis, and the specificity and effectiveness of each function strongly affect the pattern of spontaneous mutations.

spontaneous mutations occur more randomly and much less frequently.

There are several ways in which mutations can be classified; some of the common ones are

1.Direction of mutation: a. Forward mutation:(mutation from wild type allele) b.Reverse mutation:(mutation from a mutant allele to

the normal one) 2.Cause of mutation: a. Spontaneous mutation:(occur naturally without any

apparent cause)

Induced mutations:(originate due to a treatment with some physical or chemical agent)

3.Dominance relationship: a. Dominant mutation: b.Recessive mutation:(most often recessive) c.Codominant mutation:(occationally codominant) d.incompletely dominant mutation:(partially dominant) 4.Tissue of origin: a.Somatic mutation:(mutation occuring in somatic cell) Bud mutation:(somatic mutation occurs in an

axillarybud)

b.Germinal mutation:(occur in reproductive tissue) In an sexually reproducing species only germinal

mutations are transmitted to the next generation. 5.Effect on survival: a.Lethal mutations:(kill all the individuals having them in

appropriate genotype) b.Sublethal mutations:(kill most of the individuals) c.subvital mutations:(kill some of the individuals) d.vital mutations:(do not affect survival) e.supervital mutations:(increase the survival)

6.Type of affected: a.Morphological/visible mutations:(mutation that alters

mutant allele is visually detected) b.Biochemical mutations:(mutations prevent the

production of a biochemical by the organism) 7.Quantum of morphological effect produced: a.Macro mutations:(produce large enough changes in the

phenotype detected without any confusion) e.g:short legged mutation in sheep.

b.Micro mutations:(quantitative characters generally produce small morphological effects,special techniques required for their detection)

8.Cytological basis: a.chromosomal mutations:(detectable changes in either

chromosome number or structure) e.g.,bar eye mutation in drosophila b.gene mutations/point mutations:(produced by

alternations in the base sequencens of genes) c.cytoplasmic or plasmagene mutations:(changes in

mitochondria or chloroplast DNA) e.g.,cytoplasmic male sterility in maize, sorghum.

9.Molecular basis: a.base substitution:(single base in a DNA molecule is

replaced by another base) b.deletion mutation:(one or more bases are deleted or lost

from a gene) c.addition mutation:(insertion of one or more bases in a

gene) 10.Type of amino acid replacement: a.missense mutation:(replacement of a single amino acid

of the respective polypeptides by another amino acid) They are produced by base substitutions.

b.nonsense mutation:(a codon does not code for any amino acid)

c.frameshift mutations:(all the amino acids of the polypeptide chain located beyond the site of mutation are substituted)

Frameshift mutations are produced due to deletion or addition of bases.

1.Errors during DNA replication. 2.mutagenic effects of the natural environments of

organisms. 3.transposons and incertion sequences. 4.methylation followed by spontaneous deamination of

DNA bases, esp cytocine. 5.some of the solar radiations are surely mutagenic. Both eukaryotic and prokaryotic genomes contain some

transposable elements.e.g.,incersion sequences(IS),transposons,etc.

The rate of spontaneous mutations is very low. It generally ranges between 10⁻⁵to10⁻⁷ /gene

/generation for prokaryotes. 10⁻⁴ to 10⁻⁶/gene/generation for eukaryotes. Spontaneous mutation rates per generation are

comparatively higher in eukaryotes than in prokaryotes. But some eukaryotic genes are highly stable and mutate

at a very low rate. e.g.,the waxy locus in maize. Estimated mutatin rates in man appear to be much higher

than those for other eukaryotes.

The rate of forward mutations to yellow body colour and to brown eye colour in drosophila are estimated as 1.2 х10⁻⁴ and 3 х 10⁻⁵,respectively.

Mutations produced due to the treatment with either a chemical or physical agent.

The agents capable of inducing mutations are known as mutagens.

The capacity of an agent for inducing mutations is termed as mutagenic property.

Induced mutations are useful in two different ways: 1.in genetic and biochemical studies. 2.in crop improvement.

The process of induced mutations through treatment with a mutagen is known as mutagenesis.

The exploitation of induced mutations for crop improvement is called mutation breeding.

Some potent chemical mutagens may produce mutations at the rate of more than 1% per gene per generation in bacteria and bacteriophages.

This represents a more than 10,000-fold increase in the mutation rate.

A large variety of microbial mutants have been isolated and studied intensively by microbiologists . some important ones:

1.AUXOTROPHIC MUTANTS

2.RESISTANT MUTANTS

3.CRYPTIC MUTANTS

4.CONDITIONALLY EXPRESSED MUTANTS

5.ANTIGENIC MUTNTS

6.METABOLIC MUTANT

7.REGULATORY MUTANTS

8.TEMPARATURE-SENSITIVE MUTANTS.

1.AUXOTROPHIC MUTANTS: Those that are nutritionally deficient ,i.e., they are unable

to synthesize essential metabolites or growth factors(amino acid ,purine ,pyramidine,vit.,).

2.RESISTANT MUTANTS: It’s exhibit an increased tolerance to inhibitory agents,

particularly antibiotics and phases. Microbes may develop such mutants spontaneously

through a range of mechanisms. 3.CRYPTIC MUTANTS: Those that have lost a specific function but retain the

intracellular activities.e.g,.loss of permease.

4.CONDITIONALLY EXPRESSED MUTANTS: These mutants remain as wild type phenotype under one

set of conditions. 5.ANTIGENIC MUTNTS: It’s show a change in the surface structure and

composition of the microbial cell. 6.METABOLIC MUTANT: These exhibit altered metabolic ability particularly the

fermentation ability of decreased or increased capacity to produce some end-product.

7.REGULATORY MUTANTS: Mutation affects either regulatory region of the promoter

gene or the activity of a regulatory protein. Mutants of Bacillus subtilis are grossly deficient in the

enzymes. 8.TEMPARATURE-SENSITIVE MUTANTS: These will grow at one temparature but not at another.

most of them are heat-sensitive ,some how-ever , are cold sensitive.

Stress Survival

Adaptive mutation

Transformation of Escherichia coliwith the F' plasmid containing the lac operon. The operon possesses a + 1 frameshift, so it is unable to express genes for lactose catabolism. Following specific cultivation conditions with lactose containing medium, the lactose operon on some of the plasmids revert to lac+.

The recombination-dependent mutation model for adaptive mutations of the lactose operon on the F' plasmid. The “leaky” +1 frameshift of the lac operon provides enough energy from lactose catabolism to initiate replication of the plasmid. (A) Persistent nicks in the plasmid cause the replication fork to collapse and leave exposed dsDNA. (B) Exposed dsDNA initiates DNA repair with RecA, which inserts a ssDNA segment into the dsDNA. (C) Replication of the DNA by the low fidelity DNA polymerase IV gives an increased potential of replication errors leading to numerous mutations.

Genomic rearrangement resulting from activity of an indigenous ISelement. (A) IS5 inserts between the CRP-binding site and the cstA gene.(B) IS5 then undergoes inversion with IS5D, which is already locatedupstream of an ABC-type transporter operon. (C) The inversion causesthe regions between IS5 and IS5D to invert. Hence, the ABC-typetransporter operon is now positioned near the CRP-binding site andbecomes activated.

Confirmational change of the carboxyesterase. The esterase (left) can hydrolyze carboxy esters, but the confirmation specificity of the enzyme’s catalytic site does not allow hydrolysis of other polymers, such as nylon. Point mutations in the enzymes’ gene can cause a conformational alteration of the enzyme’s catalytic site so that specificity is reduced (right). This reduced specificity now allows the enzyme to hydrolyze a wider variety of oligomers, including the linear polymer, nylon-6.

Bacteria frequently develop mutations that enable them to survive and adapt to a variety of environmental conditions.

However, most of these mutations can be classified as a form of antagonistic pleiotropy.

Bacteria face a variety of environmental conditions and stressful situations.

If the environmental conditions change, the mutation usually becomes less beneficial and perhaps even detrimental.

most bacteria need the ability to rapidly adapt to ever changing environments and food sources.