Module 3 Biology PDF

Lecture 1 DNA, Genes, and Chromosomes 1) DNA stores genetic information a) Encodes for proteins that provide structure and do much of the work of the cell 2) Genetic information is stored in genes 3) Turning on a gene, or gene expression, ultimately causes an effect on the organi...

Lecture 1 DNA, Genes, and Chromosomes 1) DNA stores genetic information a) Encodes for proteins that provide structure and do much of the work of the cell 2) Genetic information is stored in genes 3) Turning on a gene, or gene expression, ultimately causes an effect on the organism 4) Gene regulation controls when the process should occur DNA = Deoxyribonucleic Acid - A Casual Relationship between structure and functions exists in many molecules - The structure of DNA allows it to store and transmit genetic information Griffith’s experiment - Virulent bacteria → make mouse sick with pneumonia - Nonvirulent bacteria → mouse not sick - Killed virulent bacteria → mouse not sick - Killed virulent + nonvirulent bacteria → mouse dies of pneumonia Griffith concluded that nonvirulent bacteria can become virulent; form of inheritance; plasmids → component DNA that is transferred one to another → nonvirulent bacteria takes up the DNA from virulent and become virulent bacteria - Some type of molecule in the killed debris carried genetic information for virulence \\ Avery, Macleod, and Mccarty’s experiment - DNase of virulent bacteria extract are being destroyed and therefore, no virulent bacteria is made in the final solution - DNA was responsible for the genetic exchange of information; made up of gene and chromosome Hypothesis is based on previous knowledge, scientific intuition, educated guess that would allow us to set up a new experience; always has to be measurable Control Group → we know what the end goal will be; for comparisons to predict to the experiments DNA is built from nucleotides 1) Deoxyribose sugar 2) Phosphate group 3) Base (A,C,T, and G) a) Purines: (Adenine A and Guanine G); b) Pyrimidines: (Thymine T and Cytosine C); i) Pairings: AT (2HB) and CG (3HB) Sugar + Base = Nucleoside Sugar + Base + Phosphate = Nucleotide or Nucleoside monophosphate Phosphodiester bonds 5’ end 5’-AGCT-3’ Always read from 5 - 3 3’ end iClicker Question Why aren’t nucleosides incorporated into DNA? a) There are no phosphates to make the phosphodiester bonds b) The bases are not fully assembled c) The sugar is not in the right form d) The peptide bonds don’t form The sugar phosphate backbones wind around the outside of the molecule and the bases point inward - Forms a helical structure - Bases lock tgt the two strands The two strands in the backbone are antiparallel, which means that they run in opposite direction - The antiparallel allows the backbone being stabilized the ability for the base pairs to interact with each other - A%=T% ; G%=C% iClicker Question In the DNA of certain bacterial cells, 16% of the nucleotides are adenine. What are the percentages of the other nucleotides in the bacterial DNA? a) 34% thymine, 34%guanine, 16% Cytosine b) 34% uracil, 16%guanine, 16% Cytosine c) 16% thymine, 34%guanine, 34% Cytosine d) 16% thymine, 16%guanine, 34% Cytosine Hydrogen bonding and base stacking contribute to the stability of DNA - Hydrophobic interaction between the plane of the bases that also contribute to the strength of holding the shape DNA is compacted in cell - DNA is cells must be compacted - Too long to fit within the diameter of cell - Each DNA molecule in a eukaryotic cell forms one chromosome - Linear DNA molecule - DNA is packaged with proteins - Proteins known as histones interact with DNA without regard to sequence - Histone-DNA complexes further compact DNA = chromatin - Histones + DNA = chromatin = form of DNA that makes up chromosomes within our cells Lecture 2 The Histone proteins are rich in positively charged amino acids (lysine and arginine). Why do you think this is important for their function? - Because the nucleotides have negatively charged backbone which can be bound by the positively histones and condense down to chromosomes Each DNA molecule in our cells form one linear chromosome - We have 23 pairs of chromosomes - 22 autosomes and 1 sex chromosome - In each pair, one is from biological mother and one is from biological father Chromosomes - 2 strands of DNA hooked with centromere - Each strand is double stranded dna molecule; 2 sister chromatids held tgt; 2 linear DNA molecule Each DNA molecule in our cells forms one linear chromosome DNA = Deoxyribonucleic Acid - A Casual Relationship between structure and functions exists in many molecules - The structure of DNA allows it to store and transmit genetic information - Need to make copies of DNA = replication - This replication needs to be exact - Errors = mutation in DNA DNA Replication - Taking parental strand and separate to create 2 complementary base pair strands → daughter strands - Parental strand serves as a template Rosalind Franklin – photograph 51 - Discovered that bases are pointed inside - DNA is double helix S phase - Homologous chromosomes duplicate to create sister chromatids - 2 linear DNA to 4 linear DNA Hypothetical Models for Replication - Semiconservative - The new DNA duplex consists of one old strand (parental) and one new strand (daughter) - Conservative (WRONG) - The new DNA duplex consists of two newly synthesized daughter strands, leaving the parental duplex intact The Meselsonstahl experiment: DNA replication is semiconservative Eukaryotic Dna Replication is also semiconservative Replication occurs 5”to 3” - Daughter - Carried out by DNA polymerase - Needs 3’ OH to start attaching nucleotide; RNA primase RNA primase (an RNA polymerase) lays down an RNA primer - Creates the little primer 5’-3’ DNA polymerase extends the RNA primer. It cannot begin a new strand on its own but can only elongate the end of an existing piece of DNA or RNA. Continuous/Discontinuous Replication - Continuous/ Leading strand – bottom strand - Because the parent/template strand starts from 3’-5’; only need 1 primer RNA primer - Discontinuous/ lagging strand – top strand - parent/template strand starts from 5’-3’ - Uses a lot of RNA primer – synthesized by primase - Okazaki fragments : tiny strands of replicated DNA (daughter strand) - DNA polymerase pulls out the RNA primer and replaces it with DNA nucleotides - DNA ligase then joins the two DNA fragments Proteins involved in replication Helicase – unwinds the DNA into two separate strands Topoisomerase II – prevent twisting; undoing the knotting before helicase Single Strand binding proteins – keeping the two parental strands apart DNA synthesis of both strands occurs at the same time and rate - DNA polymerase complexes of each strand stay in contact with each other and build strands at same rate - Requires lagging strand be looped around to maintain the contacts between the polymerase complexes Proofreading – by DNA polymerase What could happen if the proofreading mechanism does not spot a mistake? - Mutation Replication origin iClicker To the right of the dotted line, where is the lagging strand synthesized? a) Top strand b) Bottom strand c) Both strands Both leading and lagging occurs on one parental strand For bacteria – replication of circular DNA Linear DNA after each round of replication End replication problem issue - RNA primase takes up itself so by the end, it will be shorter and shorter - Telomeres – caps = chromosome ends - To not lose important genetic information What Exactly is the problem, again? In lagging strand synthesis, multiple RNA primers but they never gets to the end - The daughter lagging strand is being shortened - Telomeres – repeated sequences that are maintained by telomerase - It allows the shortened 3’ end of the template strand to be restored by the addition of more telomere repeats – extend parental strand Telomeres can report on your age - Telomerase activity differs between different cells → stem cells have fully active telomerase while in adult cells in many parts of the body it is inactive - When telomeres shorten to about 100 copies of telomere repeats, cell division will stop = aging - Healing more slowly - 50-100 times for replication of telomeres to occur before they shut down the telomere process Cancer cells seem to show increased activity of telomerase. Why do you think this might be the case? Wanted to be copied – ability to continue cell division even with mutations Lecture 3 DNA Manipulation - Using the knowledge of replication to manipulate DNA - PCR (polymerase chain reaction) - Amplify a region or gene using the same process the DNA polymerase uses to replicate - Gel Electrophoresis - Separate the DNA fragments by size and visualize them (ex: through UV light) PCR (polymerase chain reaction) Denaturation - Taking two strands of DNA where the Target sequence reside - Usually involves high heat to separate - Close to boiling ~98 C degrees Annealing - Flood the sequence with primer (DNA sequences) that we manufactured that are complementary to one and the other strand - Forward (A) and reverse primers (B) - Anneal to the target region; allow the primers to get to the binding spots before the two strands come back tgt Extension - DNA polymerase added will then use the primer added from PCR to create the two pieces of amplified DNA Runs around 30 cycles of PCR everytime - 2 n copies of template sequence where n is the number of cycles after each application cycle After Denature, due to hot boiling temperature, proteins are usually dead by that time. So to prevent that and allow for 2nd, 3rd,4th.. Time of PCR to happen thermus aquaticus. A bacteria that loves hot environment and grow - Taking the DNA polymerase from thermus aquaticus into PCR - Will allow polymerase to cycle 30 rounds when denaturing Run the products of PCR on agarose (Gel electrophoresis) - Add DNA to the wells on electrophoresis gel and allow to move in electrofield - DNA that has the negative phosphate backbone will move to the positive electrode - Agarose allows us to see how fast a large/small piece can move through - Large pieces of DNA stop short - Small pieces of DNA migrate further iClicker Question Using PCR, a student hopes to amplify a sequence of known DNA from a genomic DNA sample. The two primers are both designed to pair with sequences that have a 40% guanine-cytosine (GC) content. Using a particular set of denaturation temperatures, annealing temperatures, and extension temperatures, the experiment produces not the expected single DNA fragment of a single known size but a set of fragments of different sizes. Which of the factors could account for these results? a) The denaturation temperature was too high b) The annealing temperature was too high c) The sequences of the primers were not specific enough d) The extension temperature was too low DNA Editing Many techniques exist that allow us to specifically modify the sequence of a gene or piece of DNA - E.g. reverse a mutation in a gene to restore normal function = DNA editing - CRISPR/Cas 9 CRISPR/Cas 9 - Enzyme that identifies as being a bad thing (virus), goes up to the bad thing and cuts it - CAS 9 – cutting - CRISPR – RNA that acts as the memory and find the virus that’s trying to enter the body - CRISPR uses the RNA template to identify virus (have memory of virus) DNA Editing - CRISPR and Cas 9 work tgt - Exonuclease will increase the cut space (due to virus being cut out of DNA) - Thenediting template DNA will be added to the gap between the target DNA CRISPR has the potential to alter our genes in any way we want - In lab, CRISPR has been used to fix the mutations in DNA that cause diseases like cystic fibrosis - Monkeys were used to test CRISPR/Cas9 DNA editing - Created with changes in 2 genes - Editing human embryos is possible - Possible to have off-target effect risk DNA editing in cystic fibrosis Cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride ion channel - Chloride ions will move into the lumen alongside with water - Forskolin – chemical that are activated to allow CFTR to function properly - Mutation in CFTR leading to cystic fibrosis (CFTR F508Del) - Leading to dehydration; improper hydration of airways and intestines - Organoids – a assembly of cells; miniature copy outside of body - Cell edited with CRISPR to correct mutation of cystic fibrosis → F508Del-Corrected clone (S1-c1 and S1-c2) Genetic information in DNA directs the activities in a cell Our cells have been manipulating DNA forever - The Central Dogma - Transcription and Translation - Exceptions: viruses and prions Lecture 4 The Central Dogma Carry information in the form of DNA - Transcribe into mRNA - Translate mRNA → Protein Transcription → make mRNA RNA vs DNA (Ribose and Deoxyribose) 1) Ribose has a hydroxyl group where deoxyribose has a hydrogen group 2) CG pairing remains the same, AT pairing changes; T → U (Uracil) a) Uracil has a hydrogen (-H) group and Thymine has a methyl group (CH3) 3) RNA has a nucleoside triphosphate at the 5’ template end; DNA has a nucleoside monophosphate at the 5’ template end 4) DNA double strand; RNA single strand RNA World Hypothesis - Many scientists believe the first nucleic acids were RNA molecules - RNA is involved in many cellular processes, including all the steps of the central dogma - RNA also has enzymatic properties - DNA is used by cells because it is more stable than RNA molecules Transcription - DNA serves as the template for RNA production by the cell - Although transcription occurs in different places in prokaryotes and eukaryotes, the process is similar in both types of cells - Take the 2 parental strands apart; picking only the sequence of one of the parental strand as template strand to transcribe mRNA Template vs Nontemplate strands Genes can be transcribed using either parental strand as long as they have the right orientation The sequence of an RNA transcript is: 5’AUGUCGUAAG 3’ What is the sequence of the template strand DNA? Template strand: 3’ TACAGCATTC 5’ Nontemplate strand: 5’ ATGTCGTAAG 3’ Initiation and Termination of Transcription - DNA molecule usually contains many genes - Close to the promoter region on template strand is where the transcription begin - Terminator is where the transcription stops  - Transcription is initiated at a promoter sequence and ends at a terminator sequence. The transcription is always synthesized in a 5’ to 3’ direction. RNA polymerase and associated proteins bind to the DNA duplex at the promoter. iClicker question Genes can be coded on either strand of DNA. Consider the transcribed regions of two genes (gene A and gene B). The arrows represent the regions of the genes that are transcribed and the direction in which transcription occurs. For Gene A, which strand will be the template strand? a) The top strand as shown in diagram b) The bottom strand as shown in the diagram c) Neither strand Initiation of Transcription - Transcription factors and RNA polymerase bind double-stranded DNA at promoter sequences - Promoter sequences are conserved DNA sequences - Promoter - Certain sequences (T and A) are repeated several times and are called TATA box Initiation and termination of transcription Many eukaryotic promoters contain a sequence similar to TATAAAA = TATA box First nucleotide to be transcibed usually positioned about 25 base pairs from the TATA box - Meaning RNA are getting started to be assembled 20-25 pairs away from the TATA box in the direction of RNA building (5’ - 3’) Prokaryotic cells dont have TATA box but have something else similar that acts the same way (-10 or -35 elements) - - indicate that they transcribe behind the promoter site Promoter recognition in eukaryotes is complex Enhancers are located in, near, or some distance from a gene - Necessary to begin RNA transcription A general transcription factor known as TATA-box binding protein (TBP) interacts with the TATA box region of the promoter during transcription initiation - Enhancer sequences are binded by transcriptional activator proteins - Help us begin transcription Mediator complex is best friends with RNA polymerase - Bring RNA polymerase to start RNA transcription Looping of DNA is important to start transcription Elongation – adding on RNA till u reach terminator RNA replication reaction is really similar to DNA replication - Only difference is T turns into U RNA polymerase in Prokaryotes →similar to Eukaryotes 1) DNA enters RNA polymerase → unwinds 2) RNA polymerase start letting RNA nucleotides in to transcribe RNA 3) Allow the already transcribed DNA leave the polymerase Primary Transcript in Prokaryotes Does Not have to wait till the transcription process to end to hop into the translation process Primary transcript = mRNA Primary Transcript in Eukaryotes Transcript and Translation occur in separate compartments Eukaryotic transcripts undergo processing prior to translation = modifying Modifying - Adding 5’cap (7-methylguanosine cap) - Adding Polyadenylation = Poly (A) tail to 3’ end [around ~250A] - Protect the mRNA species; contribute to stability - Introns are being spliced out (non coding sequences) and remainder exons are connected tgt (contain meaningful sequence) - Spliceosome composed of RNA and proteins is responsible for splicing - Splicing occurs first by cleaving off from the 5’ intron and creating a lariat; lariat then break down into individual nucleotides; exons (5’ and 3’) join together - Alternative splicing: when some exons are missed and not being able to be included in the final mRNA - Create diversity When a region of bacterial DNA that codes for a protein is put into a human cell between a promoter and a terminator, the human cell makes the functional bacterial protein. When we reverse the experiment (human DNA into bacterial cells), the correct protein is often NOT made. Why? 1) Compartmentalization problem. RNA transcription and translation can both occur at the same time but DNA cannot. 2) Bacteria would not be able to process the DNA as it does not have modification like human Translation and Protein Structure Proteins are diverse and versatile Proteins are made up of building blocks called amino acids - Amino group + Alpha carbon + carboxyl group + R-group The exact sequence of amino acids will determine each protein’s shape and function How are proteins made and folded into a specific shape in order to be functional? Hydrophobic amino acids – tend to be buried in the interior folds of protein Hydrophilic amino acids – like to be water - Have polar, basic and acidic iClicker Question A folding domain of a polypeptide chain has a primary structure containing five phenylalanine residues, where F represents the side chain of phenylalanine. Consider the possible folding orientations. a) Figure A b) Figure B Special amino acids (Glycine, Proline, Cysteine) - Small and flexible - Creates kinks - Forms bridges within and between proteins = disulfide bonds Glycine – small and flexible Proline – disrupts continue chain Cysteine – disulfide bonds - Connecting 2 separate proteins into 1 Lecture 5 Formation of peptide (in protein) 2 amino acids →connecting through 1 carboxyl group and 1 amino group resulting in 1 H2O as waste (dehydration reaction) - Peptide bond holds the 2 amino acids tgt Proteins have directionality like nucleic acids and are often labeled with a N terminus and C terminus. What does this notation mean? N terminus → amino terminus, beginning of the polypeptide C terminus → carboxyl terminus, the free carboxyl group from the last peptide, end of polypeptide N terminus → C terminus just like how u read DNA 5’ → 3’ Some nomenclature: Protein = polypeptide Amino acid = residue No matter what the function is of a polypeptide, its ability to carry out that function will be dictated by its 3-dimensional shape Amino Acid Structure 1) The primary structure is the sequence of amino acids a) Simply the linear structure of amino acids from N terminus to C terminus b) EX: alanine-methionine-alanine-methionine or A-M-A-M or Ala-Met-Ala-Met 2) The secondary structure results from interactions of nearby amino acids a) Two ways of organizing: Alpha helix or beta sheets b) Occur between the amino acid functional groups, not the R groups c) Alpha Helix i) Each carbonyl group in the backbone forms a hydrogen bond with an amide group four amino acids away (1-4) ii) The polypeptide chain is twisted tightly in a right-handed coil iii) Cannot have bulky R groups d) Beta sheets i) Layer the polypeptide chain like a rope then bend it backward to allow strands to form hydrogen bonds with itself (1-37)/ (16-57) etc..  ii) Adjacent strands run in the same direction (parallel) or in opposite directions (antiparallel) iii) Hydrogen bonds form between carbonyl groups in one polypeptide and amide groups in a different part of the polypeptide 3) The tertiary structure is the 3-dimensional shape of a polypeptide a) Our body usually only go up to tertiary structure b) Tertiary structure of a protein is its three-dimensional shape, usually made of several secondary structure elements c) Determined by the spatial distribution of hydrophilic and hydrophobic R groups along the molecule as well as by chemical bonds and interactions that form between the R groups i) Where hydrophobic is buried inside and hydrophilic is on the outside ii) Disulfide bonds might form to lock the structure between cysteine 4) The quaternary structure results from interactions of polypeptide subunits a) Possible structures: i) Enzyme consists of two identical polypeptide subunits ii) Alpha Helix interact with Beta sheets (1) EX: Hemoglobin: 2 alpha and 2 beta Primary structure → shape Shape = function - Some might new help of slowly folded proteins that bind newly made proteins and help them fold called Chaperone Protein function can be altered by the environment of the protein - Chaperone will help protect against denaturation - Most proteins don't necessary need chaperone How is the sequence of amino acids specified? Translation!!! Require mRNA, ribosome, transfer RNA(tRNA), aminoacyl tRNA synthetases, initiation,elongation and release factors Ribosomes - Made up of proteins and ribosomal RNAs - Ribosome moves down the mRNA from 5’ to 3’ and reads individual codons to incorporated the appropriate amino acids - Large subunit and small subunit - Large: used when landed on first AUG; Exit site, Peptidyl site, Aminoacyl site - Small: initial that bound to first tRNA - Start Codon: AUG Three functional sites of the Ribosome (in large subunit) - A site: accepted the aminoacyl tRNA - P site: peptide bond formation occurs - E site: where tRNA exits the ribosome AUG = Methionine - Translation begins at the start codon UAA/ UAG/UGA= Stop codon Initiation in prokaryotes The initiation complex can be found at multiple internal sequences where a Shine-Dalgarno sequence is found. Polycistronic mRNA = codes for multiple protein - Produces multiple protein from 1 mRNA Have to land on Shine-Dalgarno sequence to start translation (contains AUG) Initiation in Eukaryotes Monocistronic mRNA – only 1 protein from 1 mRNA Initiation in eukaryotes begins at the 5’ cap, the first AUG is the start codon Transfer RNA (tRNA) - Each tRNA has the nucleotide CCA at its 3’ end, and the 3’ hydroxyl of the A is the attachment site for the amino acid tRNA synthetases - Binds to 1 uncharged tRNA and its corresponding amino acid - Then the enzyme attaches the amino acid to the 3’ end of the tRNA Aminoacyl tRNA synthetases - Responsible to detect codon and link correct amino acids to tRNA iClicker Refer to the mRNA sequence: 5’ – AUGAGACUUACCGAA – 3’. What would the anticodon look like if the second nucleotide of the fourth codon was mutated to U. a) 3’ –TAG –5’ b) 3’ –UAG– 5’ c) 3’ –AAC –5’ d) 3’ –AUG– 5’ Many amino acids are specified by more than one codon Process of Translation 1) Initiation a) Recruit small ribosomal subunit and tRNAmet (having UAC codon complementary to AUG) and scan the mRNA for an AUG codon – binds to the 5’ cap b) Large ribosomal subunit joins in when AUG is found and initiation factors are released 2) Elongation a) Bring in new tRNA in the A site and when matched, peptide bond between the start and newly matched amino acid will be linked b) Uncharged tRNA now moved to the E site, allow new tRNA to enter A site c) Polypeptide transfer the amino acid on tRNA in A site. 3) Termination a) Encounter a stop codon → release factor enters A site b) Polypeptide is released from the tRNA in the P site c) Ribosome dissociates Protein Families - Protein are similar enough in sequence and structure that can be grouped tgt - Many members share small regions of 3-D structure = folding domains Protein SOrting - Not only form protein in cytosol, but also the ER Lecture 6 Cap-dependent and cap-independent translation Eukaryotic initiation factor – 4G (eIF-4G) - The initiation factor that start scanning from the 5’ cap to find AUG , where protein synthesis begins Virus Poliovirus - Encodes poliovirus protein that chops up eIF-4G protein needed for scanning from cap - Results in no translation of capped mRNAs → cannot scan AUG - No more host cell proteins → only poliovirus protein will be translated How does poliovirus mRNA translate? - Have IRES that acts like 5’cap - Poliovirus proteins are made because they are not capped - They have internal ribosome entry site (IRES) Regulation of Gene Expression in Eukaryotes The Human body - 200 major cell types - All share same genome - Look and function differently from one another - Because each cell type expresses different sets of genes Eukaryotic gene regulation - How cells control gene expression - Can occur at many levels 1) Epigenetic control a) Modifying DNA or genome by chemically changing something on the DNA b) Attaching chemical groups onto histone tails → compaction/loosen DNA i) Chemical modification of histone tails repositions nucleosomes and exposes stretches of DNA to the transcription machinery (1) Tightening histone holding onto DNA → less access for transcription (2) Loosening histone onto DNA → allow more access for transcription Histone tail modification - Patterns of modifications = histone code = affects chromatin structure and gene transcription - Methylation → tightening - Setilation → loosening Methylation of DNA - DNA can be methylated - When Cytosine is beside Guanine, cytosine will be methylated - Lightly methylated → transcription occur  - Heavily methylated → transcription does not occur; because proteins are recruited to remodel chromatin, modify histone tails (compacted the region of the genome) Epigenetic effect - Changes nothing about the DNA sequence itself or promoter - Only added chemical modification to the existing base pair - Can be passed on to next generation if it is heavily methylated gene in the gamete Methylation can be reversed Development of an embryo Totipotent = fertilized human egg - give rise to all cell types in a complete organism + extraembryonic structures + placental cells - Yes embryonic tissue Pluripotent = cells of the inner cell mass that are embryonic stem cells - give rise to any of cell of the body but cannot be entire human body - No extraembryonic tissue - Inner cell mass of blastocyst - Methylation of specific genes happened Multipotent = cells of the germ layers - Form a limited number of types of specialized cell - Methylation of specific genes happened Are all capable of differentiating into different cell types and sustain life *Genes have to be silenced in order to be differentiated - Through acetyl group taken away and methyl group added to silence Induced pluripotent stem (iPS) cells as therapy iClicker question Which cell, or cell type, has the greatest developmental potential? a) Gastrula b) Inner cell mass c) Fertilized egg (zygote) d) Ectoderm e) Nerve cell Regulation of an entire chromosome Why would you inactivate a whole X chromosome? - To compensate the dosage - Dosage compensation = the level of expression of X-linked genes is the same in both sexes X-Activation The Xist gene is transcribed and Xist RNA binds with the X-chromosome inactivation center (XIC) - Xist continues to be transcribed and later on code the entire X-chromosome - Presence of Xist RNA triggers DNA methylation and other associated with reduced transcriptional activity - *Xist RNA and Xist gene only works when there is XX chromosome 2) Transcriptional control - Enhance or silencer sequences - Enhance bind activator proteins - Silencer binds repressor proteins → inhibits RNA polymerase 3) Translational control - Species that can inhibit translation: siRNA (small RNA) and miRNA (micro RNA) - Associated with RISC complex and target certain mRNA - siRNA pair with target mRNA, (pair perfectly) leading to RISC complex to chew up target mRNA - miRNA (doesnt pair perfectly) pair to target mRNA, signal to RISC complex and inhibit target mRNA Translational regulation: proteins that bind UTRs - Having elements on the mRNA that can be bind by other proteins - UTR (untranslated region) - mRNA sequence recognized by binding protein can block ribosome Perfect Translational control using the UTR binding site Iron is transported to transferrin and bind it to transferrin receptor → cells having iron Too much iron? → stores in protein called ferritin 5’ IRE - BP= iron responsive element binding protein Lecture 7 iClicker Question 1 Which of the cartoons best represents ferritin mRNA under LOW IRON conditions? A Under low iron conditions. The IRE-BP is active and bound to the IRE. When bound to the ITR, translation is blocked = no ferritin storage protein is made - Because there is little iron, there is no need for iron storage protein (ferritin) - Block translation for storage protein iClicker Question 2 The transferrin receptor gene encodes an IRE within the 3’ UTR of the mRNA. RNA instability elements are found within the IRE. Under high iron conditions: a) The IRE-BP would block access to the instability elements, increasing the lifespan of the mRNA and allowing more transferrin receptor to be made b) The IRE-BP would not block access to the instability elements, decreasing the lifespan of the mRNA and limiting protein production c) The IRE-BP with iron would make the instability elements ineffective How is Genetic Material Inherited? Cell Division - The process by which cells make more cells - Cell division occurs for: - Growth - Cell replacement - Healing - Reproduction Cell Cycle (eukaryotes) M phase (mitosis and cytokinesis) Interphase: G1, S, G2 phase (G0) G1 phase (preparation for DNA synthesis) - Preparing for duplication of DNA S phase (DNA Synthesis) - Duplicate DNA G2 phase (preparation for mitosis and cytokinesis) - Physically expand in size G0 phase (resting phase for some cell) iClicker Question 3 The chromosomes in the circled region could both be chromosomes from the same parent a) True b) False Centromeres/chromatids iClicker Question 4 Which circle region contains two identical pieces of double-stranded DNA? a) b) c) d) e) None are identical Mitosis Stage 1) Prophase 2) Prometaphase 3) Metaphase 4) Anaphase 5) Telophase + Cytokinesis Mitotic cell division - Results in 2 identical daughter cells - Each daughter cell contains the same number of chromosomes as the parent cell iClicker Question 5 Colchicine is a drug that blocks the assembly of microtubules. If dividing cells are treated with colchicine, at what stage of mitosis would you predict the arrest of cell division to occur? a) Prophase b) Metaphase c) Late anaphase d) Telophase iClicker Question 6 A skin cell in G2 of interphase has ____ as much DNA as it had in G1. a) Half b) Twice c) One-fourth d) Four times Meiotic cell division - Results in 4 daughter cells - Each cell contains half number of chromosomes as the parent - Each daughter cell is genetically unique Prophase I - Homologous chromosomes align next to each other, forming the bivalent - Crossing over happens → chiasma - Increases genetic variation because it results in combination of alleles not seen in the parental chromosome - Results in 2 sets of recombinant and 2 sets of non-recombinant chromatids Prometaphase I and Metaphase I - Meiotic spindles attach to kinetochores on chromosomes, leading to metaphase I where bivalents move to the center of the nucleus Anaphase I and Telophase I - Homologous chromosomes have separated, resulting cells are haploid Results in 2 haploid Meiosis II - Sister chromatids will separate, resulting in gametes - Meiosis II is often called the equational division - Often called the equational division Results in 4 haploids Cytoplasmic divisions Male: 4 gamete (sperm cells) Female: 1 gamete(oocyte), 3 polar bodies - The single oocyte receives most of the cytosol  - Nondisjunction Nondisjunction → not separating Parthenogenesis = reproduction from an egg without fertilization Lecture 8 How is genetic material inherited? Part 2 Transmission genetics - Each of us has his or her own personal genome - It differs from all that have existed before and from all the will come after - Transmission genetics = manner in which genetic differences among individuals are passed from generation to generation Pea Plant crossing - Pea flowers have sperm-and egg-producing structures that allow for self-fertilization to occur - Mendel removed sperm-producing structures in order to ensure that only his intended cross would happen Yellow vs Green Seed Traits P1 (parental generation) – true breeding strains that are crossed - The parents - True breeding – homozygous F1 generation - The outcome of P1 crossing - Usually dominant Reciprocal crosses – doesn’t matter which is male or female - Because they all receive same outcome Allele – different forms of a gene – hereditary factors The combination of alleles in an individual is its genotype and the expression of the trait is its phenotype F2 generation - From the self fertilization of F1 generation - 3:1 recessive trait The principle of segregation: the equal separation of alleles of a gene into different gametes; half get one allele, the other half get the other allele Zygote – fertilized egg cell What would be the genotypes and phenotypes of F1 progeny from a cross of an AA plant with an Aa plant for seed colour? Segregation of Alleles in meiosis Separating Maternal and Paternal chromosomes in meiosis I Incomplete dominance - Use superscripts to indicate the alleles - Ex: CRCW - Red + white = pink Codominance - Each allele produces a distinct phenotype that can be detected in heterozygous individuals - Blood type: A and B are codominant/ 50% type A and 50% type B - Red + white = Red + white Probability Probability of occurrence of a genotype lies between 0 and 1 - Aa x AA → probability of the genotype aa is 0 - AA x aa → probability to the genotype Aa is 1 Sometimes it is necessary to combine the probabilities of 2 or more possible outcomes of a cross 1. Multiplication rule: outcomes can occur simultaneously and the occurrence of one does not impact the likelihood of the other a. The probability of rolling double four: i. ⅙ x ⅙ = 1/36 2. Addition rule: possible outcomes cannot occur simultaneously (either of the mutually exclusive events occurring) a. The probability of rolling a seven in any combination i. 1/36 + 1/36 +1/36 + 1/36 +1/36 +1/36 = 6/36 = ⅙ Addition/Multiplication Rules Mating of Aa x Aa iClicker Question What is the probability that an individual is either Aa or AA if his or her parents are both heterozygous for the trait? a) ¼ b) ½ c) ¾ d) 1 Independent Assortment: segregation of one set of alleles of a gene pair is independent of the segregation of another set of alleles of a different gene pair Independent assortment of genes in different chromosomes reflects the fact that non-homologous chromosomes can orient in either of two ways that are equally likely Mendel’s laws 1) Principle of segregation 2) Principle of Independent Assortment a) Not all genes undergo independent assortment b) Some that are too close tgt do not assort independently Epistasis: two genes interacting affect the same trait Diagram of family history = pedigree Lecture 9 Pedigree of a dominant allele If one of the parent has an dominant affected trait → approximate half of the following pedigree will have the same trait - Affected individuals will appear in every successive generation Pedigree of a recessive allele A rare recessive trait may skip one or more generations - Affected individuals can have parents who are not affected - Affected individuals often result from mating between relatives Genetic testing - Method of identifying the genotype of an individual, who might be at risk for a certain trait Benefits - Personalized medicine/treatment, informed decisions about healthcare - Better understanding of risks, behaviours - Feeling less anxious, leading to a better quality of life Risks - Limited answers - physiological/emotional impact - Privacy concerns (life,health insurance) Uncommon inheritance patterns Human sex chromosomes - X is longer than Y Segregation of the sex chromosomes - ½ XX and ½ XY X-linked Genes F 1 are all red eyes A male with an X - linked recessive trait will have heterozygous (carrier) daughters and unaffected sons. “Crisscross” = X from Dad to daughter Then from that daughter to son “W-” stands for recessive white-eyed mutation in one x chromosome Normal chromosome separation Most XY males receive their X chromosome from their mother (for fruit flies only) Abnormal chromosomal separation: Nondisjunction ( for fruit flies only ) - XXY = female - X = male, only need Y to be fertile (XO) - XXX or OY were never observed = lethal Nondisjunction of Sex chromosomes in human X-linkage recessive mutation - Red green colorblindness - Hemophilia The overall frequency of recombinant chromosomes is a measure of the genetic distance between the two genes. Genes that are linked have a recombination frequency between 0% - 50% - 50% is due to only the inner two chromatids being recombinant with each other. Genetic map - Recombination frequency are additive - When the genes are close together, they are being inherited tgt and recombination will not occur Location of mutant gene a) Can be seen that GC has mutant (49) Inheritance of mitochondrial and chloroplast DNA Mitochondria and chloroplast have their own genomes - Code for enzymes that carry out the functions of that organelle - These genes move with the organelles during cell division - These genes are inherited independently from nuclear chromosomes Inheritance of mitochondrial DNA - All the offspring of mothers with the trait will show the trait - Males do not transmit the trait - When there is a fusion between egg and sperm in fertilization, all the sperm mitochondria is being destroyed within the egg; only maternal mitochondria remain in the fertilized egg - Bi-parental inheritance of mitochondrial DNA (WRONG) - Have a defect that results in a mutant protein. Was not able to destroy the paternal mitochondrial DNA after fertilization iClicker Question If this trait is X-linked recessive, what are the chances that these parents will produce an affected son as their new child? a) 100% b) 50% c) 25% d) 0% Lecture 10 Genomes A genome is the genetic material of a cell, organism, organelle, or virus, and its sequence is the order of bases along the DNA or (in some viruses) RNA. A genome is the genetic material transmitted from parents to offspring The number of genes in a genome and the size of a genome do not correlate well with the complexity of an organism Genome size of humans is around 3100 MB. Gene number is not a good predictor of biological complexity. Complexity actually correlates to all the stuff we talked about before. Sequence composition of the human genome 1) Coding 2) Introns 3) Retrotransposons: transpose by means of an RNA intermediate a) An RNA piece that creates a DNA copy of itself and insert somewhere in genome 4) DNA transposons: replicate and transpose via DNA replication and repair 5) Alpha satellites 6) Others 7) Untranslates 5’ and 3’ ends of mRNA *transposons – DNA that replicates and insert itself into new positions in the genome - Can be consequences; duplicated landing in the middle of coding gene which disrupts the outcome Mutation = any heritable change in the genetic material - Reproductive cells: germ-line mutations - Passed on to next generation - Non-reproductive cells = somatic mutations - Arise over the course of time of an individual How Do mutations arise? Mutations that are able to survive under the presence of antibiotics. - Mutations arise without regard for the needs of the organism - Mutation was already present prior to plating on selection media!! - Exposure to mutagens Point mutations – changes in a single nucleotide \ Silent mutation/synonymous mutation: change in nucleotide does not change amino acid missense mutation/ nonsynonymous mutation: results in change of amino acid Nonsense mutation: results in stop codon Insertion/deletion: adding or deleting codons Frameshift mutation: if insertion/deletion did not result in exact multiple of 3 nucleotide → create shift Genetic variation: we are all mutants! Genetic variation = difference in genotype = difference in phenotype Polymorphisms = common genetic differences that exist between individuals - Change in actual nucleotide in any one given part of genome Most genetic variation in population occurs in non-coding DNA - Neutral – no obvious effect Variable Number of Tandem Repeats (VNTR) - Different numbers of repeated sequences in particular locations on their chromosomes The Genetic and Environmental Basis of complex traits Complex traits are called quantitative traits that measure along a continuum with only small intervals between similar individuals Complex traits are influenced by the environment - Affect the variation in phenotype between individual Genotype VS environment Roles of environment vs genotype in individual is IMPOSSIBLE to determine - But possible for POPULATION No 1 genotype is the ideal genotype across the environment; no 1 environment is ideal to all genotype There is a regression towards the mean - Offspring move towards the mean Why is this observed? 1) Meiosis: recombination and segregation breaks up combination of genes that result in extreme phenotypes (e.g. very tall, very short) 2) Environmental effects are not inherited a) Environment affected the parent → but will not affect the offspring Heritability: proportion of the variation between individuals that can be attributed to genes alone