BIOL 311 Mid 2 Review PDF

**Topic 4** **Lethal Alleles:** - Lethality occurs when two copies of a mutant allele are inherited - If a mutant allele is lethal for homozygotes, then none of the offspring with that genotype will survive **Penetrance vs Expressivity** - Penetrance is the percent of individuals with...

**Topic 4** **Lethal Alleles:** - Lethality occurs when two copies of a mutant allele are inherited - If a mutant allele is lethal for homozygotes, then none of the offspring with that genotype will survive **Penetrance vs Expressivity** - Penetrance is the percent of individuals with a mutation that show the phenotype - Incomplete or variable penetrance is defined when some individuals with a mutant genotype will not show the mutant phenotype - For example, this disorder of bone formation which is a dominant disorder is never developed by some people - Expressivity is the differing levels that a phenotype is expressed - Variable expressivity is defined when some individuals show differing degrees of the phenotype - Polydactly in cats -- dominant trait but the affected cats can have a differing number of extra toes A chart of different ovals Description automatically generated **Why would individuals with the same mutation not show exactly the same phenotype?** - Environment - Other genes- genetic background - Subtlety of mutant phenotype -- missed classification **Interaction of genes in a pathway:** - Many gene are usually involved in the presentation of a certain phenotype - These genes are often in a pathway where one gene turns on the next - If any gene in the pathway is mutated, the phenotype is no longer wild type **Beadle and Tatum:** - Investigated the genetic control of cellular chemistry Neurospora - They found numerous mutant strains that were arginine auxotroph meaning they needed arginine to survive - Each mutation behaved as a single gene - They mapped the mutations relative to other genes and found that they map to three different loci  \ **Determining functional relationships between genes:** - Mutate a population to generate mutations in genes. Obtain many mutant lines - Perform complementation tests to determine number of genes mutated - Make double mutant lines to determine gene interactions **Complementation Test:** -  - A white paper with text and numbers Description automatically generated - We want the wild-type blue flower. So, mutant 1 and 3 as well as mutant 2 and 3 produce all blue meaning that they complement and have a mutation in different genes. However, mutant 1 and 2 produce white meaning that they have the mutation in the same gene and fail to complement. - Complement = mutant alleles in different genes - Fail to complement = mutant alleles in same gene **Double mutant interaction:** - No interaction (9:3:3:1) - Same pathway (9:7) - Working towards end product/phenotype - Gene products working in the same pathway - Functional (usually dominant) alleles are needed for both genes - Recessive epistasis (9:3:4) - Two products produce phenotype - Dominant epistasis (12:3:1) - One mutant hides the other - Epistasis is when the phenotype of a mutant allele masks the phenotype of the mutant allele of another gene - Suppressor mutations (13:3) - Two wrongs make a right - They reverse the effect of mutation in another gene, resulting in the wild type **Synthetic mutations (15:1):** - Mutations in two different genes individually don't cause a phenotype, but together result in a mutant (synthetic) - Often due to redundancy **Topic 5** **Two main themes underlying the observations on chromosomal changes:** - Karyotypes generally remain constant within a species - Most genetic imbalances result in a selective disadvantage - Related species usually have different karyotypes - Closely related species differ by a few rearrangements - Distantly related species differ by many rearrangements - Correlation between karyotypic rearrangements and speciation **Chromosomal rearrangements:**  **Origins of chromosomal rearrangements:** - Chromosome breakage can result in all classes of chromosomal rearrangements - A screenshot of a diagram Description automatically generated**\ ** **Aberrant crossing over at repeated sequences can also produce rearrangements:** - The blue arrows represent the repeated sequences and indicate their relative orientations. The repeated DNA sequences may be simple sequence repeats (SSRs) or transposable elements **The effects of chromosomal rearrangements:** - Each of these rearrangements can impact the phenotypes or even viability by affecting gene balance - Severity of the effect can depend on whether the individual is a homozygote or heterozygote for the rearranged chromosomes - In addition, these different types of changes to chromosomes can alter crossing over, affecting the fertility of individuals **Deletion loops form in the chromosomes of deletion heterozygotes:** - Recombination only occurs at homologous regions - Therefore, no recomb can occur within a deletion loop - Genetic map distance in deletion heterozygotes are inaccurate - A diagram of a wire Description automatically generated **Types of duplications:**  **Different aspects of the genome can duplicate:** - At lowest level exons duplicate or shuffle - At next level entire genes duplicate and create multigene families - At next level gene families duplicate to produce gene superfamilies - The entire genome duplicates, doubling the number of copies of every gene and gene family **Chromosome breakage can produce inversion:** - A diagram of different types of paracetamol Description automatically generated **Inversion can disrupt a gene:** -  **Inversion loops form in inversion heterozygotes:** - Formation of inversion loop allows tightest possible alignment of homologous regions - Crossing over within the inversion loop produces aberrant recombinant chromatids **Why pericentric inversion heterozygotes produce few if any recombinant progeny:** - Each recomb chromatid has a centromere, but they will be genetically unbalanced - Zygotes formed from the union of normal gametes with gametes carrying these recombs will be nonviable **Why paracentric inversion heterozygotes produce few if any recombinant progeny:** - One recomb chromatid lacks a centromere and the other has two centromeres - Zygotes formed from union of normal gametes with gametes carrying the broken dicentric chromatids will be nonviable **Translocations attach part of one chromosome to another chromosome:** - Reciprocal translocation - Two different chromosomes where each have a chromosome break - Reciprocal exchange of fragments -- each fragment replaces the fragment on the other chromosome - Robertsonian translocation - Chromosomal break occur at or near centromeres of two acrocentric chromosomes - Generates one large metacentric chromosome and one small chromosome, which is usually lost **Phenotypic effects of reciprocal translocation:** - Most of these don't affect the phenotype because they don't add or remove DNA - Abnormal phenotypes can be caused if translocation breakpoint disrupts a gene or result in altered expression of a gene - In somatic cells it can result in oncogene activation - Defects that are observed in translocation heterozygotes - Unbalanced gametes are produced = reduced fertility - Genetic map distance are altered because of pseudolinkage **In a translocation homozygote, chromosome segregate normally during meiosis I:** - If the breakpoints of a reciprocal translocation don't affect gene function, there are no genetic consequences in homozygotes **Chromosome pairing in a translocation heterozygote:** - In a trans hetero, the two haploid sets of chromosomes carry different arrangements of DNA - Chromosome pairing during prophase I of meiosis is maximized by formation of a cruciform structure **Semi sterility in a corn plant that is hetero for a reciprocal trans and/or inversion:** - Slightly less than 50% of gametes arise from alternate segregation and are viable - Unbalanced ovules resulting from adjacent-1 or adjacent-2 segregation are aborted **Topic 6** **Bacteria:** - Uses binary fission not meiosis or mitosis **A few important bacterial genetic traits:** - Prototroph vs auxotroph - Wild-type bacteria, with his+ genotype, is a prototroph because it can make histidine and can survive in MM, MM+his, and MM+arg - His- can't grow without histidine because it can't make its histidine, it's a his AuxotrophA group of circles with black text Description automatically generated - Ability to use a particular carbon source - Wild-type bacteria (lac+) grow in MM which contains lactose as the carbon. The lac+ bacteria can use the lactose to grow in MM - The mutant unable to use lactose is lac-, so it won't grow in the MM because it cannot break down lactose - It will however grow if it's given glucose - Overall, the his- mutant lacks the ability to produce an essential nutrient (histidine) and lac- mutant lack the ability to use a specific carbon source (lactose) -  - Antibiotic resistance - The wild type bacteria (str^S^) is streptomycin sensitive therefor unable to grow in MM containing streptomycin - Str^r^ is however, the mutant, resistant to streptomycin is able to grow in MM with streptomycin **Bacterial DNA exchange** - Horizontal gene transfer - Movement/exchange of genetic info without sexual reproduction or cell division - Within the same gen - Conjugation - Direct contact between bacterial cells via pilus, where DNA is transferred from the donor to the recipient - A small circular plasmid goes from one cell to another or a part of the bacterial genome - Transformation - Picking up free DNA from the environment/ a dead bacterial cell - Transduction - Virus mediates transfer of DNA from donor cell to the recipient cell - Used to determine distance of genes as well **Bacterial conjugation:** - Donor cell (F+ or Hfr) - Contains the F plasmid or integrated F factor - Can produce pili - Transfers genetic material to the recipient - Recipient cell (F+) - Lacks the F plasmid - Can't produce pili - Receives genetic material from donor - Fertility (F) factor is a plasmid that gives bacterial cell the ability to produce pili - HFr strain is a bacterial strain in which the F factor is integrated into the chromosome - What happens in Hfr x F- cross? - Almost none of the F- recipient strains are converted into F+ nor Hfr strains - The integrated F factor, in the HFr strain, drives transfer of some or all of the bacterial chromosome - The donor chromosomal fragment can recombine with the recipient chromosome - One strain is replicated and sends the F- to the other part - Terminus is last to replicate and this means we transferred the F- - Transfer starts at the origin and DNA replication+transfer begins where the F factor was integrated - Exconjugate is a cell that contains a fragment of donor DNA; has participated in conjugation - The donor chromosomal fragment can recombine with the recipient chromosome and this is what changes the gene **Interrupted mating to determine the order of bacterial genes in the chromosome:** - F plasmid can integrate at different sites and in different orientations - Different interrupted mating experiments will give different results - Gene order of entrance will indicate location and direction of F factor **Chromosome mapping based on recombination frequencies:** - Recombination in bacteria - Recombination takes place between a complete genome and an incomplete - Partial genome (exogenote) - Linear fragment is lost - Complete genome (endogenote) - Recombinant, intact circular genome - To keep circular genome intact, there must be an even number of recombination events - Liner + circular = linearization X - Remember that the closer two genes are together on the chromosome, the less likely a crossover will occur between them - A screenshot of a test Description automatically generated -  - A diagram of a line of lines Description automatically generated with medium confidence -  - A diagram of a dna molecule Description automatically generated with medium confidence - Quadruple crossovers rarely ever occur -  **Mapping the viral genome:** - Phage DNA can be circular or linear **Bacterial DNA Exchange: Transduction** - Transduction is the transfer of genetic material from a bacterial donor to a recipient by a phage - Generalized transduction is the random incorporation of bacterial DNA into phage heads - The closer the bacterial genes are in the chromosome, the more likely they are to be packaged into a phage head and be transduced together (co-transduced) - Higher co-transduction means closer the genes are - Specialized transduction - Is a process of genetic transfer in bacteria involving temperate phages - Virulent phages: immediately lyse and kill the bacterial host (lytic) - Temperate phages: integrate their DNA into the host chromosome without killing it (lysogenic, bacteria with phage integrated) - The phage integrated into the bacterial genome is called prophage - The bacteria harboring the prophage is called lysogen - Prophages sometimes become active and cause lysis of the host cell - Lambda phage DNA is Circular - Special transduction cont'd - Lysogen production - Lambda is circular and integrates into bacterial chromosome - Lysate production - Normal outlooping = phage DNA excises normally - Abnormal = phage DNA excises incorrectly by taking adjacent bacterial genes. For ex. Gal+ gene - Defective phage formation - The abnormal outlooping phage can infect a new bacterial cell, potentially transferring the bacterial genes it carries - What is the major difference between cotransduction experiments and recombination frequency experiments? - The larger the recomb freq the further apart the genes are - The larger the cotrans freq the closer together the genes are **Topic 7 and 8** **DNA Structure -- required properties:** - Must allow for accurate replication - Must contain information - Must be able to change (rarely) **Building Blocks:** - Pure as gold (purine, A and G) - Cut the py (pyrimidine, C, U, and T) - RNA has two hydroxyl groups and DNA has one **Base pairing:** - Same amount of purine nucleotides and pyrimidine - Same amount of A and T; same amount of C and G - A+T isn't necessarily equal to G+C - Most GC rich is more stable than AT **Double Helix:** - Nucleotides contain a phosphate, sugar, and base - Nucleotides form DNA strands by phosphodiester linkages (links the 3' carbon of one sugar to the 5' carbon of another via a phosphate) - The two sugar-phosphate backbones are antiparallel - DNA strands are held together by 2-3 hydrogen bonds between purine/pyrimidine - A=T and G triple bond to C **DNA is replicated in a semi conservative manner:** - Semi-conservative replication - Two DNA strands unwind from each other - Each DNA strand acts as a template for the synthesis of a new complementary strand - Results in two double helices that are identical to the original **DNA replication -- synthesis in the 5' to 3' direction:** - Replication is catalyzed by DNA polymerase - Cleaving off pyrophosphate produces energy to help drive DNA synthesis **DNA replication:** - Single strand DNA binding proteins stabilize unwound DNA - Relaxes supercoil DNA and rejoins the DNA strands - Helicase disrupts the H- bonds between strands - DNA polymerase III catalyzes DNA synthesis. Can extend a chain but can't start a chain - DNA replication is semi-discontinuous - RNA polymerase synthesizes the primer - DNA polymerase I removes the primer and fill in the gap (5' to 3') - DNA ligase joins the fragments together - Ligase catalyzes phosphodiester bond **PCR (polymerase chain reaction) method of amplifying DNA:** - Detect the presence or absence of this gene - Getting enough DNA copies to perform sequencing, enabling us to detec a mutation - Cloning and genetic engineering **Components needed to do PCR:** - Template DNA - DNA primers - DNA sequence approx. 20 nucleotides long with a free 3'-OH group - Design primers to bind in either side of your region of interest (fwd and rev primers) - Primer sequences are always written 5' to 3' - Reverse primer extends right to left on the top strand - Forward primer extends left to right on the bottom strand - dNTPs - DNA polymerase - Special DNA polymerase called Taq polymerase - Originally isolated from basteria in a hot spring in a national park - Active at high temps and remains active over multiple cycles of heating and cooling **PCR steps:** - Things needed - Isolated DNA (template) - Primers (several copies of both rev and fwd primers) - Taq polymerase - dNTPs - Steps - Denaturation - 30 sec heating at 95 degrees Celsius - Breaks apart the 2 strands of the DNA template - Annealing - 1 min at 55-66 degrees cooling - Primers bind at the appropritate locations on the template - The correct temp is determined based on the melting temps of the primers which is when 50% of primers are bound and 50% are not - Melting temp is determined by the length of primer (longer = high MT) and GC (high GC = high MT) - The annealing temp should also be about 5 degrees lower that the melting temp of the primers - Synthesis - 1 min at 72 degrees heating - Taq polymerase catalyzes elongation by adding nucleotides to the 3' end of the primer - Some things to note - The primer seqs are included in the PCR product - The fwd primer looks the same as the top strand - The rev primer is the rev complement of the top strand **DNA sequence variation:** - Both common polymorphisms and rare mutations are the result of changes in the DNA sequence - SNPs (single nucleotide polymorph) - This type of change can result in either common polymorph or rare mut - Most common type of genetic variation among humans - Single base pair differences between DN A seqs - Transition vs Transversion SNPs - Transition replaces pyrimidine with pyrimidine (PYT PYT) - Transversion replaces purine with a pyrimidine or vice versa (PUT PYT or PYT PUT) - Causes - Spontaneous DNA replication error (mismatch) - Bases can spontaneously change between different isoforms which tricks the polymerase into adding the wrong complementary base - Polymerase has proofreading ability so it removes mis-paired bases - 3' to 5' exonuclease activity property makes the polymerase work bwd to exise incorrect base at the end of the growing DNA chain - Mistakes that escape proofreading are normally corrected by other DNA repair mechanisms - Other chemical changes to a nucleotide (depurination, deamination) - Depurination is the hrydrolysis of the glycosidic bond between purine base and the sugar but the phosphodiester bonds remain intact - During replication either no complementary base is added or sometimes the apurinic site can pair with another base resulting in a mutation - Deamination is the hydrolytic removal of an amino group (C, G, and A contain amino groups) - Induced mutation - Exposure to chemical mutagens - Can replace a base or alter/damage a base - Indels (insertion -- deletion) - Nonrepetitive (includes single base pair ins or del) - STRs (one type of repetitive sequence, repeats of 2-9 nucleotides) - Present in exon, introns, regulatory regions, and non-functional DNA seqs - Have a high mutation rate - Number of alleles in STR regions is often large (20+) - Ex. Allele: TAG, Allele: TAG TAG TAG - SNP usually has 2 diff alleles max 4, but STR can have multiple different alleles (20+) - Since STRs are more polymorphic than SNPs, STRs are better able to discriminate between samples and confirm matches **Gel electrophoresis:** - Gel electrophoresis in used to check if PCR worked - It is a method to separate NDA molecules based on their size, using electrical field to move molecules through a gel matrix - Negative charge is applied to the top of the gel (where well is) - Positive charge is applied to the bottom of the gel - Small DNA travels quickly and further - Large DNA travels slow and doesn't get to far - Molecular ladder contains pieces of DNA with know sizes to help determine the sizes of your DNA samples - DNA travels at a rate that is inversely proportional to the log of its size - Similar steps as PCR **Sanger sequencing:** - Similar steps but with key differences being - Seq doesn't result in exponential amplification - Only one primer is needed - Add ddNTPs in addition to dNTPs - Original method (four separate rxns) - DNA template - DNA polymerase - Primers (fwd or rev) - dNTPs - ddNTPs in four separate rxns - Automated Sanger seq - All ddNTPs are added to the same rxn, with differently coloured fluorescent markers - Separated by capillary electrophoresis and detected by laser beam