Gene Mutations I and II Flashcards
Polymorphism meaning?
Natural variation in gene,DNA, Chromosome that has no adverse affects to the organism.
Benefit - population
Benefit evolution
natural variations in a gene, DNA sequence, or chromosome that have no adverse effects on the individual and…
occur with fairly high frequency in the general population.
Beneficial – Evolution
What is a Mutation? When occurs? Effect?
Change in genome, spontaneously, polymorphism
- Change in the genome - can occur spontaneously
- Produces genetic variation (polymorphism)
3.Gene mutation = change within a single gene
- One allele is mutated, to become another allele
4.Some mutations are point mutations, change in a single nt pair
Allele meaning?
Allele = one of a number of alternative forms of the same gene or same genetic locus
Mutation Nomenclature:
for
Bases, Deletion, Insertion, Amino Acids
5162 G –> A
5162= base position
G = orignal base
A = replacement base
Deletion: 197delAG
Insertion: 2552insT
R197G
R= Original Amino Acid
197 = AA position
G= Replacement AA
Wild type vs Mutant:
Wild-type: STANDARD GENOTYPE/PHENOTYPE
- Wild-type = found in nature or in the laboratory STOCK of the organism
Mutant: ALTERED genotype or phenotype (due TO MUTATION )
Origins of Mutations:
INDUCED VS SPONTANEOUS
Which one is the ultimate source of genetic variation? rate
- Induced = Treatment with Mutagen
- Spontaneous = Absence of mutagen
- Spontaneous mutations - ultimate source of genetic
variation
(DNA replication/repair).
- Rate is low - one cell in 10^5 - 10^8
What are Mutagens? how they effect?
Mutagens cause mutations - greater dose, more mutations
A mutagen is a CHEMICAL or PHYSICAL AGENT capable of INDUCING CHANGES in DNA called mutations
define mutagenesis?
the production of genetic mutations.
TYPE SOF MUTATIONS: POINT MUTATIONS
2? What are they?
Two main types:
- Base substitutions: one base pair is replaced by another
- Transition: replacement of a base by another base of the same chemical category (Purine ->Purine e.g. A ->G) or (Pyrimidine->Pyrimidine e.g. C ->T)
- Transversion: replacement of a base by another base of a different chemical category (Purine ->Pyrimidine e.g. A ->C) or (Pyrimidine->Purine e.g. C ->A) - Base insertions/deletions (indel): Insertion or deletion of nt pairs
Types of Base Substiutions:
Base substitutions: one base pair is replaced by another
- Transition: replacement of a base by another base of the same chemical category (Purine ->Purine e.g. A ->G) or (Pyrimidine->Pyrimidine e.g. C ->T)
- Transversion: replacement of a base by another base of a different chemical category (Purine ->Pyrimidine e.g. A ->C) or (Pyrimidine->Purine e.g. C ->A)
Types of Base Insertions/Deletions (indel):
Base insertions/deletions (indel):
Insertion or deletion of nt pairs
TYPES PF MUTATIONS = Point mutations at
***AT PROTEIN LEVEL (Protein coding part of a gene) =4
- Synonymous mutation – Codon change results in same AA (degeneracy of genetic code)
- Missense mutation (conservative/non) – Change in AA (to chemically similar
AA/chemically different AA - affects protein structure and function!!) - Nonsense mutation – Codon for one AA changed to STOP codon (premature
termination of translation – affects protein function e.g. inactive proteins) - Frameshift mutation – Indel (AA sequence downstream of mutation changed – loss of normal protein structure and function)
AMINO ACID CLASSIFICATIONS
NON POLAR -
POLAR -
ELECTRICALLY CHARGED- ACID VS BASIC
NON POLAR - Glycine, alanine, valine, leucine, isoleucine, methionine, tryptophan, phenylalanine, proline
POLAR - serine, threonine, cystine, tyrosine, asparagine, glutamine
ELECTRICALLY CHARGED-
ACID: apartic acid, Glutamic acid
BASIC: lysine, arginine, histidine
Transitions vs Transversions
TRANSITIONS
1. Purine to Purine
A - G
G - A
- PYRIMIDINE
T - C
C - T
TRANSVERSIONS
1. PURINE TO PYRAMIDINE
A-C
A-T
G-C
G-T
- PYRIMIDINE TO PURINE
C-A
C-G
T-A
T-G
Consequences of mutations within genes;
Types of Mutations at DNA level, Results at molecular level
NO MUTATION
TRANSITION OR TRANSVERSION
INSERTION OR DELETION
NO MUTATION: wild type
- Codons specify wild type protein
TRANSITION OR TRANSVERSION
- Synonymous mutation: Altered codon specifies the same A acid.
- Missense mutation (conservative):
Altered codon specifies a chemically similar amino acid - Missense Mutation (NONCONSERVATIVE):
Altered codon specifies a chemically dissimilar amino acid. - Nonsense mutation: Altered codon signals chain termination (STOP CODON)
INSERTION OR DELETION:
1. Frameshift mutation:
POINT MUTATION DO NOR ALWAYS HAVE PHENOTYPIC EFFECT:
Other than silent mutations, usually the least severe mutations in a protein coding gene result from missense mutations, but some missense mutations can be very deleterious…
Mutations can alter pre-mRNA Splicing
HOW? WHAT THE STEPS?
- Whole exon can be deleted as result of mutations in splice
signals,
e.g. Exon Skipping
2.Part of exon can be deleted as a result of mutations in normal splice signals cryptic splice site selection (sequences resembling normal splice signals)
Abnormal Splicing of b-globin
RNA in patients with b
thalassaemia (Mild to severe
anaemia)
Abnormal Splicing of b-globin
RNA in patients with b
thalassaemia (Mild to severe
anaemia)
PG 19
Different single-base mutations cause different splicing defects & disease:
WHAT ARE THEY? = 3
- Exon skipping
- Cryptic splice-site selection
- New splice site created
Wild type:
which way for Wild type and mutant?
Forward vs Reverse Mutation?
- Forward Mutation – alters WT
(Changes WT into a Mutant Phenotype)
Wildtype -> Mutant
2.Reverse Mutation – get WT
(Restores the WT phenotype)
Wildtype <- Mutant
Explain Reverse Mutations:
Exact reversion (true or genotypic) gives the wild type DNA sequence
Equivalent reversion gives a different DNA sequence, but the wild type AA sequence.
“reversion” is usually used to indicate reappearance of parental phenotype in a mutant organism caused,
Change in nt sequence to give back wild-type AA sequence, but not wild-type nt sequence…
Explain Suppressor mutation:
- “hides” the effect of another mutation.
- Distinct to exact reversion in which the mutated site changes back to original WT sequence.
- Occurs at a site distinct from the site of the original mutation.
- Double Mutant - Possesses original and suppressor mutation
but WT phenotype. - Two classes Intragenic and Intergenic
Intragenic vs Intergenic Supressor
both classes of Supressor mutation:
- Intragenic suppressor – mutation occurs in the gene
containing the original forward mutation
… Might change a second nucleotide in the same codon.
…Might also work by suppressing a frameshift mutation
- Intergenic suppressor – mutation occurs in a gene other than the one containing the original mutation that it supresses
action of an intergenic suppressor mutation,
steps
Base substitution?
changes the base of a single DNA nucleotide
transition?
Base substitution in which a purine replaces a purine or a pyrimidine replaces pyrimidine.
Transversion
base substitution in which a purine replaces pyrimidine or a pyrimidine replaces a purine
Insertion vs deletion
insertion: addition of one or more nucleotides
deletion: deletion of of one or more nucleotides
frameshift mutation:
insertion or deletion that alters the reading frame of a gene
in-frame deletion or insertion:
deletion or insertion of a multiple of 3 nucleotides that DOES NOT alter the reading frame
expanding trinucleotide repeats:
repeated sequence of 3 nucleotides (trinucleotide) in which the number of copies of the trinucleotide increases
forward vs reverse mutation:
forward mutation: changes the wild type phenotype to a mutant phenotype
reverse mutation: changes a mutant phenotype back to the wild-type phenotype
missense mutation
changes a sense codon into a different sense codon, resulting in the incorporation of a different amino acid in the protein
nonsense mutation:
changes a sense codon into a nonsense codon; causing premature termination of translation
Silent mutation:
changes a sense codon into a synonymous codon, leaving unchanged the amino acid sequence of the protein
Neutral mutation:
changes the AA sequence of a protein without altering its ability to function
Loss-of-function mutation vs Gain of function mutation:
Loss-of-function mutation: causes a complete or partial loss of function
Gain of function mutation: causes the appearance of a new trait or function or causes the appearance of a trait in inappropriate tissue or at an inappropriate time.
Lethal mutation:
causes premature death
supressor mutation:
suppresses the effect of an earlier mutation at a different site
Intragenic suppressor mutation vs Intergenic suppressor mutation:
Intragenic suppressor mutation:
supresses the effect of an earlier mutation within the SAME gene
Intergenic suppressor mutation:
suppresses the effect of an earlier mutation in ANOTHER gene
Explain Gain of function mutation:
mutant vs wild type
Protein or Gene product has enhanced function or a new
function
eg Tumour formation, mutation in gene that encodes a receptor for a growth factor, mutated receptor may stimulate
growth all the time even in absence of the growth factor
Wildtype
Inactive –> Active
Mutant
Always Active
Understanding Conditional Mutations:
Mutant phenotype only seen under CERTAIN CONDITIONS
usually associated with PROTEIN FOLDING
eg at elevated
temperatures
…eg. dominant heat-sensitive lethals of
Drosophila
Understanding Somatic and Germinal Mutation
Mutations can occur in somatic cells or in germinal
cells
- Somatic cells: cells of the body (non-reproductive cells)
- Germinal cells: give rise to gametes, so DNA is passed
to next generation
Understanding Somatic Mutation and Cancer…
Cancer cells:
- control of cell division lost
- movement of these cells through the body
Mutation in genes that control cell division can lead to cancer
Detecting mutations: WHY AND HOW?
Why?
- To understand mutation events
- To obtain mutants in a particular biological process
How?
- Use systems where a mutation will cause a change in phenotype
Detecting Recessive Mutations in Humans: 3
- Only see mutant phenotype in homozygote
- (Both alleles must be mutant in order for the mutant phenotype to be observed) - Mutation may have been present for generations before phenotype seen
- X-linked recessive mutations are expressed
more frequently in males
- (need to inherit only a single copy of the mutant allele to display the trait)
Haemophilia? Expression on X and Y Chr.s?
Haemophilia: failure in blood clotting
- Mutant allele on X chromosome (Xa)
- Males carrying mutant allele (Xa Y) express phenotype
Understanding mutation rate?
How to calculate?
Mutation Rate:
Number of mutations per unit of time (eg cell divisions or generations)
Calculation:
No. mutational events / No. cell divisions
Understanding mutation Frequency?
How to calculate:
How often a mutant is found in a population?
No. Mutants / Size of Population
Understanding Mutation Frequency
(Single Genes)
examples corn vs humans
Mutations are rare events
Corn
1 x 10^-6 to 500 x 10^-6 mutations per gamete
Humans
1 x 10^-6 mutations per gamete
Strategies to Improve
Success when Hunting for Mutants? 2
- Mutagens: Agents that increase the frequency of mutation
- Selective systems:
Methods to identify mutants easily from a large population
Understanding Selective Systems:
Methods to “select” mutants
Must be able to:
- Identify mutants clearly
- Subject a large number of individuals to
- selection (mutants are rare)
What are Biochemical Mutants?
Prototroph vs Auxotroph
Biochemical Mutants: Microorganisms
Prototroph: able to grow on a simple medium
Auxotroph: requires additional nutrients to grow
- Selection of auxotrophs by inability to grow on
minimal medium (eg mutation disrupts ability to
synthesize one or more biomolecules)
1940 Beadle and Tatum’s work with Neurospora crassa
Beadle and Tatum’s work with Neurospora crassa is a classic experiment in genetics that helped establish the “one gene, one enzyme” hypothesis. Here it is in very simple dot points:
- Organism Used: Neurospora crassa, a type of bread mold.
- Objective: To understand how genes control the production of enzymes.
-
Steps:
- They exposed Neurospora crassa to radiation to induce mutations.
- Each mutation affected a specific gene.
- They observed the effects of these mutations on the mold’s ability to grow on different nutrient media.
-
Key Finding:
- Mutations resulted in the loss of specific enzymes needed for nutrient synthesis.
- For example, one mutation prevented the mold from making an enzyme to produce a specific nutrient.
-
Conclusion:
- Each gene in the mold’s DNA coded for a specific enzyme.
- Mutations in genes led to the loss of corresponding enzymes, affecting the mold’s ability to grow on certain nutrients.
-
Hypothesis:
- “One gene, one enzyme” - Each gene controls the production of a specific enzyme.
-
Significance:
- This experiment laid the foundation for our understanding of the relationship between genes and enzymes, advancing the field of molecular biology.
In simple terms, Beadle and Tatum’s work with Neurospora crassa showed that genes are responsible for making specific enzymes, and mutations in genes can lead to the loss of these enzymes, affecting the organism’s abilities. This discovery had a significant impact on our understanding of genetics and biochemistry.
Explain Resistance selection:
E. coli resistance to bacteriophage T1
Sensitive cells are lysed, do not form colonies
(a) Resistant cells form a colony
(b) - stable, genuine
mutants
Origin of Resistant cells:
Understanding the nature of mutation, 1943: Knew
phage caused E. coli death, but also knew that a
small percentage of colonies did not die - they
became resistant & were stable & genuine mutants.
BUT, didn’t know HOW!
Did the presence of the bacteriophage T1 induce a
physiological change in the E. coli that caused
resistance?
OR
Were the mutant E. coli formed spontaneously, but
randomly in time?
Origin of resistant cells
1, Physiological change
- Random mutation
Origin of resistant cells: FLUCTUATION TEST
- “Fluctuation Test” of Luria & Delbrük, second scenario
correct. - Inoculated 20 small cultures & grew to 108 cells/mL
- At same time inoculated 1 large culture & grew to 108
cells/mL - Took 20 individual cultures & 20 samples from large
culture & plated in presence of phage. - 20 individual cultures - showed high variation
(“fluctuation”) in number of resistant colonies: from 0 to
107 per plate. - 20 samples from large culture had much less variation -
from 14 - 26 resistant colonies per plate.
ORIGIN OF RESISTANT CELLS BEST EXPLANATION:
Best explanation = mutations occurred randomly in time: early mutations gave higher number of resistant cells
(more generations of descendants).
Later mutations gave fewer resistant cells.
Hence, resistant cells are selected by environmental
agent (e.g. phage), rather than produced by it.
Can existence of mutants in a population before selection be demonstrated directly?
Replica plating technique (1952)
Grew cells on a master plate of nonselective media &
colonies transferred to a series of selective media plates
SUMMARY
- Mutations can be detected by phenotype using appropriate crosses, & pedigree analysis
- Mutations are rare
- Mutation is a random process. Any allele in any cell can mutate at any time!
- Mutagens increase the number of mutants
- Mutations are useful in research
- Selective systems aid in screening for mutants present in a population
