bio 207 8 and 9
what does radiation increase
Radiation greatly increases mutation rates in all organisms.
Pyrimidine dimer
two thymine bases covalently bind and block DNA replication.
DNA Repair Mechanisms:
Translesion Polymerases
What is the SOS system in bacteria?
The SOS system allows bacterial cells to bypass replication blocks through a mutation-prone pathway.
What does DNA polymerase η (eta) do in eukaryotes?
DNA polymerase η (eta) replaces damaged DNA with AA in eukaryotic cells to bypass replication blocks.
What do high-energy radiations (X-rays, gamma-rays, cosmic rays) do to DNA?
They remove electrons from atoms, altering the chemical structure of bases and causing double-stranded DNA breaks.
What effect does UV radiation have on DNA?
UV radiation causes pyrimidine dimer formation (usually TT, but also TC/CC), which stalls or stops DNA replication.
What is the role of the SOS system in bacteria?
he SOS system bypasses pyrimidine dimers during DNA replication, but at a high error rate.
Why would a cell want to bypass replication blocks?
To prevent cell death and survive DNA damage, even though it increases the risk of mutations. Bypassing blocks allows replication to continue under stress, though it’s error-prone.
What are double-stranded breaks in DNA?
Double-stranded breaks (DSBs) are when both strands of the DNA helix are severed, which can lead to significant damage. They can be caused by high-energy radiation (X-rays, gamma-rays) or certain chemicals and pose a risk to the integrity of the genome.
what is a historical event that lead to many somatic muatiosn in survivors
Hiroshima was destroyed by an atomic bomb inn
1945.
What were the effects of the atomic bomb's radiation on survivors?
Survivors of the initial blast suffered from radiation sickness and somatic mutations, most commonly leading to leukemia.
Were there increased germline mutations in the children of atomic bomb survivors?
Estimates showed no increased germline mutations, likely because those with the highest exposure did not survive.
What are base analogs and how do they cause mutations?
Base analogs are chemicals that resemble DNA bases in structure. When incorporated into DNA, they can lead to incorrect base pairing and cause mutations.
What are alkylating agents and give an example?
Alkylating agents donate alkyl groups (like a methyl or ethyl group) to DNA, causing mutations. An example is Ethylmethylsulfonate (EMS) and mustard gas, which was used in WWI chemical warfare (now banned).
How does deamination cause mutations?
Deamination, caused by chemicals like nitrous acid, removes an amine group from a base, altering its pairing ability and leading to mutations.
What is the effect of hydroxylamine on DNA?
Hydroxylamine adds a hydroxyl group to bases, causing changes in base-pairing that can lead to mutations.
How do oxidative reactions contribute to mutations?
Oxidative reactions, like those caused by superoxide radicals or hydrogen peroxide, can damage DNA by altering base structures, leading to mutations.
What are intercalating agents and how do they cause mutations?
Intercalating agents like proflavin, acridine orange, and ethidium bromide insert themselves between DNA bases, causing frameshift mutations during DNA replication.
What is 5-Bromouracil and how does it function as a base analog?
5-Bromouracil resembles thymine, but it has a bromine atom in place of the methyl group on the 5-carbon atom. It can cause mutations by incorrectly pairing with adenine instead of guanine during DNA replication.
What is 2-aminopurine and how does it function as a base analog?
2-Aminopurine resembles adenine but has an amino group at the 2-position. It can pair with thymine or cytosine, causing mutations by mispairing during DNA replication.
Incorporation of 5-bromouracil into a DNA
strand can lead to a replicated error, how?
if 5-bromouracil is incorporated into DNA in place of thymine, it may mispair with guanine in the next round of replication, the next replication this G pairs with C leading to permamnent muation, if it pairs with A then no replicated error occurs.
basically incorporation of bromouracil followed by mispairing leads to a TA--> CG transtion mutation
what type of muations does EMS/alkylation cause
transition mutation during DNA replication
CG--> TA
TA--> CG
what type of mutation does nitrous acid (HNO2)/deamination cause
Nitrous acid (HNO₂) causes deamination of cytosine, converting it into uracil. During DNA replication, uracil pairs with adenine instead of guanine, leading to a transition mutation (cytosine to thymine) after several rounds of replication.
CG-->TA
TA-->CG
what type of mutation does hydroxylamine (NH2OH)/hydroxylation cause
hydroxylamine (NH₂OH) adds a hydroxyl group to cytosine, converting it into hydroxycytosine. This altered base mispairs with adenine instead of guanine, leading to a transition mutation (cytosine to thymine) during DNA replication. only CG-->TA way not vice versa
C-->T
G-->A
How does EMS (ethylmethane sulfonate) cause mutations?
EMS is an alkylating agent that adds an ethyl group to normal nucleotides, resulting in mis-pairing during DNA replication. This causes point mutations, such as G→A or C→T transitions.
How do oxidative radicals affect guanine?
Oxidative radicals convert guanine into 8-oxy-7,8-dihydrodeoxyguanine, which can mispair with adenine during DNA replication, leading to G→T transversions.
What is the difference between a transversion and a transition mutation?
Transition: A mutation where a purine (A or G) is replaced by another purine, or a pyrimidine (C or T) is replaced by another pyrimidine (e.g., A→G or C→T).
Transversion: A mutation where a purine is replaced by a pyrimidine, or a pyrimidine is replaced by a purine (e.g., A→C, G→T).
How do intercalating agents like proflavin and acridine orange cause mutations?
Intercalating agents insert themselves between adjacent bases in the DNA, distorting the helix's three-dimensional structure. This leads to insertions and deletions during DNA replication, causing frameshift mutations.
How do we test chemicals to identify mutagens?
The Ames test uses his⁻ strains of bacteria to test chemicals for their ability to cause his⁻ to his⁺ mutations. Since mutagenic activity is closely linked to carcinogenic potential, the Ames test is widely used to screen chemicals for their cancer-causing ability.
How are the bacteria prepared for the Ames test?
the test uses his⁻ strains of bacteria (bacteria that cannot grow without histidine) that have been genetically modified to be sensitive to mutations.
What is the first step in the Ames test?
The first step is to expose the his⁻ bacteria to the chemical being tested, along with a plate containing minimal histidine, which allows only bacteria with mutations that restore their ability to make histidine to grow.
What happens if the chemical causes a mutation in the bacteria during the Ames test?
If the chemical causes mutations, some his⁻ bacteria will mutate to his⁺, meaning they can now make histidine and grow. The number of his⁺ colonies indicates the mutagenic potential of the chemical.
How do we determine if a chemical is a mutagen in the Ames test?
A higher number of his⁺ colonies compared to a control (untreated) sample indicates that the chemical is mutagenic and may have carcinogenic potential.
Why is the Ames test useful in cancer screening?
Since mutagenic chemicals are often carcinogenic, the Ames test serves as a fast, preliminary screening method to identify chemicals that could potentially cause cancer.
How is EMS (ethylmethane sulfonate) used in forward genetic screens?
EMS is an alkylating agent that induces point mutations in the genome. In forward genetic screens, it is used to create mutations in a population of organisms. Researchers then observe the phenotypes and use these to identify genes involved in specific biological processes or traits.
What is the goal of forward genetic screens?
Forward genetic screens aim to identify genes that contribute to a specific phenotype or trait by inducing mutations and observing the resulting changes in the organism's traits.
What is the problem researchers face after conducting a genetic screen?
After a genetic screen, researchers have a collection of new mutants, but they don’t know if the same genes have been mutated multiple times in different individuals.
What is the role of complementation tests in forward genetic screens?
Complementation tests are used to determine if mutations in different mutants affect the same gene. If two mutants with the same phenotype complement each other, they are in different genes. If they don’t complement, they are likely mutations in the same gene.
How can you identify genes necessary for learning and memory?
1. Forward Genetic Screen: Use mutagenesis (like EMS) to create mutations in a population of organisms, then observe how these mutations affect learning and memory behaviors.
2. Behavioral Assays: Test the organism for learning and memory tasks (e.g., maze learning, fear conditioning) to identify mutants with impaired learning.
3.Gene Identification: Once mutants are found, use genetic mapping or next-generation sequencing to identify the specific genes involved.
4.Complementation Tests: If multiple mutants share similar learning defects, perform complementation tests to determine whether mutations are in the same or different genes.
5.Gene Expression Analysis: Analyze the expression of candidate genes in the brain during learning-related tasks to confirm their involvement in memory processes.
How can you identify genes necessary for learning using EMS mutagenesis?
1.Feed EMS to male flies and allow them to mate.
2.Take the progeny and establish heterozygous +/m stocks (~200 progeny).
3.Test homozygous m/m flies for learning ability.
4.Keep stocks where homozygotes show poor learning (e.g., 10% of stocks = ~20 lines).
5.Two extreme possibilities:
Extreme I: 20 mutations in 20 different genes.
Extreme II: 20 mutations in the same gene.
How do complementation tests work to determine if mutations are in the same gene?
purpose: Determine if two mutations (e.g., m1, m2, ...) are alleles of the same gene.
Procedure:
1.Cross m1/+, m2/+, and m3/+ heterozygotes with wild-type alleles.
2.All heterozygotes (e.g., m1/+, m2/+, m3/+) show wild-type phenotypes (normal learners).
3.Homozygotes (m1/m1, m2/m2, m3/m3) show similar poor learning phenotypes.
If two mutations produce normal phenotypes when crossed, they are likely in different genes (complementation). If they don’t, they are likely mutations in the same gene.
Do mutations m1 and m2 occur in the same gene?
1.Cross the parental m1/m1 and m2/m2 mutants.
2.F1 progeny: Test for learning ability.
3.If F1 progeny are still poor learners, this indicates that m1/m2 do not complement, meaning they are likely mutations in the same gene.
Conclusion: m1/m2 mutations fail to complement, suggesting they are alleles of one gene.
Do mutations m1 and m3 occur in the same gene?
1.Cross the parental m1/m1 and m3/m3 mutants.
2.F1 progeny: Test for learning ability.
3.If F1 progeny are normal learners, this indicates that m1/+ and +/m3 complement each other.
Conclusion: Since the mutant phenotypes complement, m1 and m3 are mutations in two different genes, not alleles of the same gene.
How do complementation groups help identify mutations in the same or different genes?
1.Test crosses are done between multiple mutant lines (e.g., m1, m2, m3, ... m6).
2.The results show whether the mutations complement (normal learners) or fail to complement (poor learners).
3.Based on the cross results, you group mutants into complementation groups:
-Complementation group I: m1, m2, m5 (mutations in the same gene).
-Complementation group II: m3 (mutation in a different gene).
-Complementation group III: m4, m6 (mutations in another gene).
Conclusion: There are 3 complementation groups, indicating mutations in 3 different genes.
What would be the likely hair color of an F1 offspring from a cross between Solomon Island blonde and Swedish blonde hair types?
-Solomon Island blonde has TYRP1- and KITLG mutations.
-wedish blonde has TYRP1- and KITLG+.
The F1 would likely NOT have blonde hair because of the genetic interactions between TYRP1 and KITLG.
F1 offspring could have brown or black hair, depending on the influence of other genes involved in hair color.
How can rescue by complementation help identify which mutation (Gene A or Gene B) causes poor learning in m3?
Problem: m3 has mutations in both Gene A and Gene B, and both cause non-functional products.
Solution: Perform complementation tests to rescue one mutation at a time.
-Cross m3 with a strain that has a functional Gene A or Gene B.
-If the m3 mutation is rescued by a functional copy of Gene A, the poor learning phenotype is likely due to the Gene B mutation.
-If m3 is rescued by a functional copy of Gene B, the poor learning phenotype is likely due to the Gene A mutation.
Conclusion: This allows you to identify which gene mutation is responsible for the poor learning phenotype.
How can rescue by complementation using genetic engineering identify which gene mutation causes poor learning in m3?
1.Introduce a wild-type copy of Gene A into the m3 fly strain.
2.Test transgenic flies for learning ability:
-If m3 flies become normal learners, the mutation in Gene A is responsible for poor learning.
-If m3 flies are still poor learners, the mutation in Gene A is NOT responsible.
3.Next, perform a rescue complementation experiment with a wild-type copy of Gene B to test if Gene B is responsible for the poor learning phenotype.
4.The gene responsible for poor learning is identified based on which rescue experiment restores normal learning.
how do transposable elements work
if there are staggered cuts in the target dna, transposable element can insert itself into the dna, the staggered cuts leave short single stranded pieces of DNA, the replication of this single stranded DNA creates the flanking direct repeats
What are transposable elements?
Transposable elements are parasitic DNA sequences that can move within the genome, often causing mutations.
What is transposition?
Transposition is the movement of transposable elements from one location to another within the genome.
What are the key features of transposable elements?
(2)
Flanking direct repeats: Short sequences that are repeated on both sides of the transposon.
Terminal inverted repeats: Sequences that are reversed and complementary at the ends of the transposon.
How can transposition occur?
Transposition can occur through DNA or an RNA intermediate.
What is replicative transposition?
In replicative transposition, a new copy of the transposable element inserts into a new location, while the old copy remains at the original site.
What is nonreplicative transposition?
In nonreplicative transposition, the old copy excises from its original site and moves to a new site, leaving no copy behind.
What is RNA intermediate transposition?
RNA intermediate transposition requires reverse transcription to convert the RNA back into DNA before integrating into the target site.
What are the two types of intermediates involved in transposition?
DNA or an RNA intermediate.
What happens during replicative transposition?
A new copy of the transposable element inserts into a new location, and the old copy remains at the original site.
What happens during nonreplicative transposition?
The old copy excises from its original site and moves to a new site, leaving no copy behind.
What is involved in RNA intermediate transposition?
RNA intermediate transposition requires reverse transcription to convert the RNA into DNA before it integrates into the target site.
What percentage of the human genome is made up of transposable element sequences?
About 45% of the human genome comprises sequences related to transposable elements, mostly retrotransposons.
How do transposons cause mutations?
Transposons cause mutations by:
Inserting into another gene, disrupting its function.
Promoting chromosomal rearrangements (e.g., inversions, deletions).
Moving genes (deleting and inserting) as they move within the genome.
how can transposition lead to deletion
pairing by looping and crossing over between two transposable elements oriented in the same direction leads to deletion
how can transposable elements lead to inversion
pairing by bending and crossing over betwen two transposable elements oreinted opposite direction leads to inversion
How do transposons contribute to chromosomal rearrangements?
Transposons can cause chromosomal rearrangements by:
Inserting at new locations, disrupting the normal structure of chromosomes.
Promoting inversions, deletions, or duplications in the genome as they move.
how can transpositon lead to one chromosme with deletion and one chromosome with duplication
misalignment and uneqal exchnage between transposable elements located on sister chromatids
What are insertion sequences like IS1 in bacteria?
Insertion sequences (e.g., IS1) are simple transposable elements found in bacteria. They consist of:
Short DNA sequences that can move around within the genome.
transposase gene, then Typically include 23bp terminal inverted repeats and 9bp flanking direct repeats.
They do not carry additional genes, only the genes required for transposition.
What is the structure of Tn10, a composite transposon in bacteria?
Tn10 has the following structure:
Two IS elements (IS10) at the ends: These are insertion sequences that mediate transposition.
Central region: Contains genes, such as antibiotic resistance genes (e.g., tetracycline resistance).
The IS10 elements are identical or nearly identical, enabling the movement of the central region between different genomic locations.
flanking direct repeat seuqences on the outside
Who was the first to discover transposable elements?
Barbara McClintock was the first to discover transposable elements in the 1940s while studying maize (corn). She identified that certain genes could move around within the genome, causing changes in the expression of other genes.
What are Ac and Ds in maize?
Ac (Activator) and Ds (Dissociation) are transposable elements in maize discovered by Barbara McClintock.
Ac is an autonomous element, meaning it can move on its own because it encodes the necessary transposase enzyme.
Ds is a non-autonomous element, meaning it cannot move on its own but relies on Ac for transposition.
How does transposition result in variegated maize kernels?
Transposition causes variegated maize kernels by disrupting the C (color) gene in maize.
Ac element produces transposase which stimulates transposition of a Ds element in the C allele. this disrupts its pigment producing function, resulting cells have genotype Ctc and are colorless
Variegation occurs because some kernels have the C gene active (producing color), while others have the gene disrupted by the transposon, leading to a lighter color or no color at all.
how can theri be mosaic tranpostion
an Ac element produces transposase which stimulates further transpositon of the DS element in some cells. As DS transposases, it leaves the C alle restoring alleles function. A cell in which Ds has transposed out of the C allele will produce pigment generating spots of color in an otherwise colorless kernel
What are the characteristics of Class I (retrotransposons)?
Long terminal direct repeats and short flanking direct repeats at the target site.
Contains a reverse-transcriptase gene (and sometimes other genes).
Transposes through an RNA intermediate.
Examples: Ty (yeast), copia (Drosophila), Alu (human).
What are the characteristics of Class II (DNA transposons)?
Short terminal inverted repeats and short flanking direct repeats at the target site.
Contains a transposase gene (and sometimes other genes).
Transposes through DNA (either replicative or nonreplicative).
Examples: IS1 (E. coli), Tn3 (E. coli), Ac and Ds (corn), P elements (Drosophila).
What are alleles?
Alleles are different forms of a gene that exist at a specific locus (location) on a chromosome.
What is a wild type allele?
A wild type allele is the most commonly found allele in a population and typically represents the normal or standard version of a gene.
What is a variant or mutant allele?
A variant or mutant allele is different from the wild type allele. It may:
Not adversely affect the gene product.
Not result in a detectable phenotype (physical expression).
Why is purple considered the wild type in certain populations?
In certain populations, the most common allele for color (such as in flowers) is purple, which is considered the wild type because it is the most frequently observed color.
What does it mean if an organism is homozygous for an allele?
An organism is homozygous for an allele if it has identical alleles for that gene on both homologous chromosomes.
What does it mean if an organism is heterozygous for an allele?
An organism is heterozygous for an allele if it has one wild type allele and one mutant allele for a given gene.
What is an allelic series or multiple alleles?
An allelic series or multiple alleles refers to the set of known mutant alleles for a gene plus its wild type allele.
What is a wild type allele?
A wild type allele is the most common allele in a population and is considered the functionally normal version of the gene.
What are homozygotes?
Homozygotes are cells/organisms that have identical alleles for a gene of interest.
Example: FC/FC or fc/fc, where both alleles are the same on homologous chromosomes.
What are heterozygotes?
Heterozygotes are cells/organisms that have one wild type allele and one mutant allele for a gene of interest.
Example: FC/fc, where one allele is wild type (FC) and the other is a mutant (fc).
What does hemizygous mean?
Hemizygous refers to a situation where a cell/organism has only one copy of a gene, locus, or chromosomal region.
What is an example of hemizygosity due to deletion?
In a deletion scenario, the corresponding gene/locus/region is deleted on the homologous chromosome, leaving only one copy of the gene in the organism.
What is an example of hemizygosity for genes on the X or Y chromosomes?
In XY individuals (males), most genes on the X or Y chromosomes are hemizygous, meaning they only have one copy of the gene because the other chromosome is missing a second copy (i.e., males have one X chromosome and one Y chromosome).
What does dominant mean in the context of alleles?
A dominant allele is one that, in heterozygous individuals (with one copy of the allele), masks the effect of the other allele. The phenotype of these individuals is the same as those that are homozygous for the dominant allele.
What does recessive mean in the context of alleles?
A recessive allele only shows its phenotype when an individual is homozygous for that allele. In heterozygotes, the recessive allele's effect is masked by the dominant allele.
What is complete dominance?
In complete dominance, one allele completely masks the effect of the other allele in heterozygotes. The phenotype of a heterozygote is indistinguishable from a homozygous dominant individual.
How do we represent dominant and recessive alleles in notation?
Dominant alleles are typically represented by uppercase letters (e.g., FC), and recessive alleles are represented by lowercase letters (e.g., fc).
What is the relationship between the FC and fc alleles in the Flower Colour gene?
The FC allele (dominant) causes a purple flower color, and the fc allele (recessive) causes a different color. Because FC/fc and FC/FC both result in purple flowers, the FC allele is dominant over the fc allele.
What does heteroallelic mean?
Heteroallelic refers to an individual that has two different mutant alleles for the same gene, one on each chromosome.
What are the alleles in a heteroallelic organism?
In a heteroallelic organism, the alleles are different mutant alleles for a given gene. For example, fc1 and fc2 are two different mutant alleles of the Flower Colour gene, resulting in a heteroallelic combination of fc1/fc2.
What is an example of a heteroallelic individual?
An example of a heteroallelic individual would be one with fc1/fc2 alleles for the Flower Colour gene, where fc1 and fc2 are both mutant alleles different from the wild type FC allele.
What is incomplete dominance (or semi-dominance)?
Incomplete dominance occurs when the alleles of a gene do not exhibit a simple dominance/recessive relationship. Instead, the heterozygote shows an intermediate (or blended) phenotype between the two homozygous parents.
What is an example of incomplete dominance?
An example of incomplete dominance is the flower color in Four-o'clock plants:
CRed/CRed = Red petals (homozygous)
CWhite/CWhite = White petals (homozygous)
CRed/CWhite = Pink petals (heterozygous, intermediate phenotype).
What phenotype do heterozygotes exhibit in incomplete dominance?
In incomplete dominance, heterozygotes show an intermediate or blended phenotype between the two homozygous parents. For example, in the Four-o'clock plants, the heterozygote CRed/CWhite has pink petals, which is a blend of red and white.
How do heterozygotes differ from homozygotes in terms of phenotype in incomplete dominance?
In incomplete dominance, heterozygotes have an intermediate phenotype, while homozygotes express the full trait of one allele. For example, CRed/CRed produces red petals, while CWhite/CWhite produces white petals, but CRed/CWhite produces pink petals.
What is codominance?
Codominance occurs when heterozygotes express the phenotypes of both alleles simultaneously, rather than showing an intermediate phenotype. Both traits are fully expressed.