* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Meiosis - DigitalWebb.com
RNA interference wikipedia , lookup
Cancer epigenetics wikipedia , lookup
Gel electrophoresis of nucleic acids wikipedia , lookup
DNA damage theory of aging wikipedia , lookup
Nutriepigenomics wikipedia , lookup
Transcription factor wikipedia , lookup
X-inactivation wikipedia , lookup
Dominance (genetics) wikipedia , lookup
DNA vaccination wikipedia , lookup
Messenger RNA wikipedia , lookup
Human genome wikipedia , lookup
DNA polymerase wikipedia , lookup
Molecular cloning wikipedia , lookup
Designer baby wikipedia , lookup
No-SCAR (Scarless Cas9 Assisted Recombineering) Genome Editing wikipedia , lookup
Site-specific recombinase technology wikipedia , lookup
Nucleic acid double helix wikipedia , lookup
Epigenomics wikipedia , lookup
Cell-free fetal DNA wikipedia , lookup
Short interspersed nuclear elements (SINEs) wikipedia , lookup
DNA supercoil wikipedia , lookup
Extrachromosomal DNA wikipedia , lookup
Genetic code wikipedia , lookup
Cre-Lox recombination wikipedia , lookup
Polyadenylation wikipedia , lookup
Genome editing wikipedia , lookup
Vectors in gene therapy wikipedia , lookup
History of genetic engineering wikipedia , lookup
Epigenetics of human development wikipedia , lookup
Nucleic acid tertiary structure wikipedia , lookup
RNA silencing wikipedia , lookup
Epitranscriptome wikipedia , lookup
Artificial gene synthesis wikipedia , lookup
Non-coding DNA wikipedia , lookup
Microevolution wikipedia , lookup
History of RNA biology wikipedia , lookup
Point mutation wikipedia , lookup
Therapeutic gene modulation wikipedia , lookup
Nucleic acid analogue wikipedia , lookup
Non-coding RNA wikipedia , lookup
Created by Julia Hsu Levy – Version 1.5 Chapter 17: Meiosis Meiosis is a process of generating haploid cells (gametic cells), cells with only one set of chromosomes. In humans, meiosis occurs only in the ovaries or testes. Meiosis allows for sexual reproduction, that is the exchange of chromosomes from two organisms. Various types of sexual reproductive life cycles: 1. Animals 2. Most fungi and some alga 3. Plants and some alga 78 Created by Julia Hsu Levy – Version 1.5 79 Created by Julia Hsu Levy – Version 1.5 Interphase I: DNA synthesis occurs. Each chromosome becomes replicated. Meiosis I: 1. Prophase I: During synapsis, homologous chromosomes pair together in tetrads. Portions where the homologous chromosomes are held close together (chiasmata, pl. chiasma) can switch places. o A phenomenon called crossing over takes place. Non-sister chromatids are crossed to hold homologous pairs together, but sometimes, portions of the chromosomes swap places. 2. Metaphase I: Homologous chromosome pairs align along the metaphase plate. Kinetochore microtubules attach, extending from the centrosomes. 3. Anaphase I: Homologous chromosomes separate and move towards opposite poles. 4. Telophase I and cytokinesis: Cleavage furrows or cell-wall formation occur to form two daughter cells. `Meiosis II 1. Prophase II: spindle apparatus forms; chromosome pairs move towards metaphase plate 2. Metaphase II: Centromeres align on metaphase plate; kinetochore fibers attach 3. Anaphase II: sister chromatids separate; individual chromosomes move towards outer pole 4. Telophase II and cytokinesis: cleavage furrow or cell-wall created; four daughter haploid cells result How does meiosis contribute to genetic variation? 1. Independent assortment: The homologous chromosomes can align in either direction at metaphase I. 2. Crossing over: Non-sister chromatids “exchange” DNA information. 80 Created by Julia Hsu Levy – Version 1.5 Chapter 18: Genetics Gregor Mendel, an Austrian monk, had too much time on his hands. In 1843, he entered an Augustinian monastery, where after years of studying religion, he studied peas. From 1851-1853. Mendel studied at the University of Vienna with the physicist Doppler, who encouraged that mathematics be used to study natural phenomena. Mendel also studied with Unger, a famous botanist. Mendel chose peas for their ease of use, rapid growth, and distinct traits. His experiments began with true-breeding plants. He hybridized two true-breeding plants and charted the results of each character. Typical monohybrid cross: looked at one character and checked for trait variations in offspring P-generation: true-breeding plants F1-generation: first filial generation or hybrid generation F2-generation: second filial generation or offspring of hybrid generation The initial idea was that hybrids should be a “mix” of the true-breeding parents. This notion was termed blending. Guess for F1: Purple flowers (x) white flowers pale purple flowers Result for F1: Purple flowers (x) white flowers purple flowers Result for F2 F1 purple flowers (x) F1 purple flowers 705 purple & 224 white 3:1 ratio What Mendel termed “heritable factor” we now call genes. Mendelian genetics: 1. Alternative versions of genes (different alleles) account for variations in inherited characters. 2. For each character, an organism inherits two alleles, one from each parent. 3. If the two alleles differ, then one, the dominant allele, is fully expressed in the organism’s appearance; the other, the recessive allele, has no noticeable effect on the organism’s appearance. 81 Created by Julia Hsu Levy – Version 1.5 4. The two alleles fro each character segregate during gamete production. “law of segregation” Homozygous: having identical alleles Homozygous recessive (rr) Homozygous dominant (RR) Heterozygous: having two different alleles (Rr) Dihybrid cross-tests: observing heritable patterns of two characters Regular Mendelian genetics follows basic rules of inheritance. The presence of at least 1 dominant allele always produces the dominant trait. The presence of 2 recessive alleles always produces the recessive trait. Autosomal conditions: genes found on non-sex chromosomes (humans #1-44) Sex-linked conditions: genes found on sex-chromosomes (humans #45-46) Incomplete dominance: The heterozygous genotype has a phenotype that appears to be the blend of the two allelic forms. Codominance: The heterozygous genotype has a phenotype of both allelic forms. 1. MN blood groups M and N are molecules found on red blood cells. can be MM, MN, or NN 2. Tay-Sachs disease: a disease that prevents production of enzymes to metabolize gangliosides (brain lipids) and accumulation leads to brain degeneration 82 Created by Julia Hsu Levy – Version 1.5 A person with Tt genotype can detect the enzyme deficiency and be fully function. This means a person who is heterozygous also shows incomplete dominance biochemically. A person with Tt genotype can produce half-functioning enzymes and half nonfunctioning enzymes. This means a person who is heterozygous also shows codominance molecularly. Multiple alleles: More than 2 alleles exist to create the phenotype. Pleiotropy: genes that affect multiple phenotypic traits Epistasis: gene at one locus that alters the phenotype of another gene at a neighboring locus Quantitative characters: characters that vary on a continuum, rather than an either/or situation (as Mendel found in peas) o Usually results from polygenic inheritance (multiple genes affect one phenotype) o Example: skin coloration, height 83 Created by Julia Hsu Levy – Version 1.5 Characters are often multifactorial: requires many proteins to create one phenotype * * * Pedigrees * * * Pedigrees: a way to show genetic history in blood relatives Widow’s peak = dominant Straight hair line = recessive * * * Disorders * * * Disorders are any non-productive phenotypes. Disorders are not evenly distributed among the various ethnic groups because of the geography (thus genetic) isolation. Recessive disorders: harmful phenotypes from recessive alleles Affected individuals: homozygous recessive (rr) Carriers: heterozygous (Rr) Normal: homozygous dominant and heterozygous (RR and Rr) 84 Created by Julia Hsu Levy – Version 1.5 Example: cystic fibrosis (1:2500 European descendants affected; 1:25 carrier) The chloride ion channels are not fully functional in homozygous recessive individuals. They are not capable of transporting chloride ions between cells or into the extracellular fluid. Because of increased (chloride ion), the mucus coat on certain cells becomes unusually thick and sticky. Mucus buildup leads to increase infections because increased mucus levels disable some antiboties in the body. Example: Tay-Sachs disease Example: Sickle-cell anemia (1:10 African descendants carriers) Affected: homozygous recessive individuals Carriers with some evolutionary advantage: heterozygous Normal: homozygous dominant Dominant disorders: harmful phenotypes from dominant allele Lethal dominant alleles are less common than recessive alleles in the population. Why? Example: achondroplasia (type of dwarfism) Example: Huntington’s disease Affects individuals ages 35-45 Degenerates nervous system Chromosome #4 may carry Huntington’s disease on a locus near the tipe. 85 Created by Julia Hsu Levy – Version 1.5 Ways to detect disorders: 1. using Mendelian crosses to show possible outcomes of diseases 2. using pedigrees 3. carrier recognition: determining whether parents are heterozygous carriers 4. fetal testing amniocentsis: Around the 14-16th week of pregnancy, doctors remove about 10 mL of amniotic fluid. They can culture cells and karyotype the chromosomes or detect for the presence of any chemicals. Chorionic villi sampling: Within 24 hours of extracting fetal tissue, cells are cultured. Karyotypes can be 86 Created by Julia Hsu Levy – Version 1.5 made of the chromosomes. Ultrasound: uses sound waves to produce images Fetoscopy: scope inserted into the uterus that transmits light Karyotypes can reveal sex of offspring and any possibility of nondisjunction. * * * The Mathematics of Genetic Probability * * * Punnett squares: brute-force way of solving genetics problems The purpose of using a Punnett square is to figure out the probability of two organisms interbreeding to produce ____ trait. Monohybrid test crosses can reveal the parental genotypes. Purple flower (x) White flower PP or Pp pp Two possible outcomes: p p PP x pp P Pp Pp P Pp Pp 100% purple, Pp p p Pp x pp P Pp Pp p pp pp 50% purple (Pp); 50% (pp) 87 Created by Julia Hsu Levy – Version 1.5 Common monohybrid ratios: Parental Genotypes AA x aa Aa x Aa Aa x aa aa x aa Offspring outcome 100% heterozygous genotype 100% dominant phenotype 25% AA, 50% Aa, 25% aa 1:2:1 genotype 3:1 phenotype 50% Aa, 50% aa 1:1 genotype 1:1 phenotype 100% recessive genotype 100% recessive phenotype MULTIPLICATION RULE: determines the probability (chance) that two independent events will occur together Example 1: What is the probability that is I toss two coins, both will be heads if the odds for heads are 1 out of 2 (½)? ½x½=¼ Example 2: A plant that is Yy genotype is allowed to self-pollinate (Yy x Yy). What chance does the offpsring have for being yy? Because the law of independent assortment Yy will segregate into separate gametes. Each parent has a ½ chance of donating a y gamete. ½x½=¼ Example 3: A plant that is Yy is crossed with a yy plant. What chances does the offspring have for being Yy? Yy parent has a ½ chance of donating y gamete. yy parent has 2/2 chances of donating y gamete. ½x1=½ Example 4: A plant has a genotype YyRr. If this plant were allowed to self-pollinate, what is the probability that the offspring will be YYRR? The parent has a ¼ chance of donating any of these gametes: YR, Yr, yR, yr. ¼ chance of YR from sperm ¼ chance of YR from ovum ¼ x ¼ = 1/16 88 Created by Julia Hsu Levy – Version 1.5 ADDITION RULE: determines the probability of events that occur in two of more ways Example: Yy sperm is fertilized with a Yy egg. What is the probability that the resulting offspring will be Yy? ½ Y from sperm and ½ y from egg ¼ probability ½ Y from egg and ½ y from sperm ¼ probability ¼+¼=½ Solving trihybrid crosses: PpYyRr x Ppyyrr What is the probability that the offspring would have at least two recessive alleles? ppyyRr, ppYyrr, Ppyyrr, PPyyrr, ppyyrr ppyyRr ¼ pp (x) ½ yy (x) ½ Rr = 1/16 Pp x Pp ¼ pp Yy x yy ½ yy Rr x rr ½ Rr ppYyrr ? Ppyyrr ? PPyyrr ? ppyyrr ? Add all the probabilities together. 3/8 offspring will have at least two recessive phenotypes. Practice problem: AaBbRr x AaBbRr What are the probabilities that the offspring will display all dominant phenotypes? 89 Created by Julia Hsu Levy – Version 1.5 Chapter 19: Gene expression * * * Transcription * * * Transcription is the process of creating an RNA (ribonucleic acid) strand from a DNA template. DNA is a double helical structure, but only one of the strands “codes” for the RNA. Transcription is slightly different in eukaryotes and prokaryotes. Phases of transcription: 1. Initiation 2. Elongation (propagation) 3. Termination PROKARYOTIC TRANSCRIPTION: 1. Initiation: process of “beginning” transcription This is the rate limiting step. If initiation does not take place, transcription cannot take place. RNA polymerase is the enzyme that must bind to DNA RNA polymerase is a holoenzyme – made of up the core enzyme (two alpha and two beta subunits) and the sigma factor. holoenzyme The core enzyme has high affinity for DNA, but the sigma factor is not added to the core enzyme until transcription is ready to occur. RNA polymerase must bind to the promoter region. The promoter region is usually a sequence of many thymine and adenine nucleotides (TATAAT) about 10 nucleotides before the start of transcription. TATA box or Pribnov box - The TATA box is said to have consensus (tightly bound to RNA polymerase). 90 Created by Julia Hsu Levy – Version 1.5 In front of the TATA box, about 35 nucleotides from the start of transcription, is another consensus nucleotide sequence. Together the TATA box and the consensus sequence form the promoter region. Promoter Operator downstream What happens if you mutate the promoter region? o DNA topoisomerase I “unwinds” the DNA by introducing negative supercoils. DNA gyrase also introduces negative supercoils. By this, they relax DNA to form an open complex. Once the DNA is opened, the core enzyme latches onto the DNA. The core enzyme slides upstream and down stream of the DNA but cannot transcribe the DNA. This is called promoter scanning. Not until the sigma factor binds to the core enzyme does transcription begins. Depending on how tightly the holoenzyme binds to the promoter region (strength) directly correlates with how efficient transcription will be. 2. Elongation: process of adding nucleotides to make an RNA strand (only single stranded) RNA polymerase (holoenzyme) begins transcription adding A-U, C-G. A = adenine U = uracil C = cytosine G = guanine RNA polymerase unwinds the DNA to create an RNA replication fork. Problems: RNA polymerase cannot “fall off” of the DNA strand in the middle of transcription. To ensure tight binding, NusA protein is used to help lock-in the tight binding. RNA polymerase must continually unwind at the head of transcription and rewind the DNA after nucleotides are wound. 3. Termination: a process of ending transcription 91 Created by Julia Hsu Levy – Version 1.5 Protein-independent: hair pin loops (stem loops, inverted repeats of G and C) Protein-dependent: Rho-factor Rho is a protein that binds to the RNA and travels behind the RNA polymerase. Once rho catches up to the RNA polymerase, termination takes place. 92 Created by Julia Hsu Levy – Version 1.5 EUKARYOTIC TRANSCRIPTION: Eukaryotes have three different RNA polymerases: 1. RNA pol I: rRNA 2. RNA pol II: mRNA 3. RNA pol III cytoplasmic and small nuclear RNA Eukaryotes also have more than 4 subunits in the core enzyme. Transcription takes place in the nucleus. Because DNA in eukaryotes is bound around protein histones (DNA + histone = nucleosomes), for transcription to take place, the protein histones must be disassembled or denatured. Transcription in the nucleus produces precursor RNA. Not until RNA processing is mRNA (messenger RNA) created and exported through nuclear pores to the cytoplasm. 1. Initiation: The TATA box is about 30 nucleotides upstream of the start. There is a consensus GC rich nucleotide sequence at 40 and 110 nucleotides upstream of start on DNA. If the consensus is CT rich, then the sequence is fungi. There are sequences called enhancers further upstream of the GC or CT rich sequences. The RNA polymerase holoenzyme binds to the promoter region (TATA box). A TATA box binding protein (TBP) is often present. 2. Elongation: similar to prokaryotes, nucleotides added 3. Termination: similar to prokaryotes RNA processing: The RNA is long and has a non-coding sequence on the 3’ end called the trailer. There are introns which are noncoding sequences interspersed with the coding RNA. The RNA made is very unstable. The initial RNA transcript is called heterogeneous nuclear RNA (hnRNA). It is easily degraded. 93 Created by Julia Hsu Levy – Version 1.5 A 5’-cap is added, and also a 3’-polyA tail is added. CAP: guanyltransferase adds GTP molecules to add a guanine to the 5’ end of the RNA. The guanine becomes methylated (add –CH3). The mRNA becomes heavily methylated to stabilize the RNA. Poly-A tail: polyadenylation (adding adenine nucleotides to the 3’ end) Small nuclear ribonucleoprotein particles (snRNP or SNURP) add the adenine nucleotides. RNA can self-splice. It will remove the RNA introns automatically. snRNP are involved in intron removal in complexes called spliceosome. In eukaryotes, the RNA is “processed.” Introns are removed Exons are joined. 5’-cap is added. Poly-A tail is added. There are two genes that encode two specific RNA strands that fold to become ribosomal RNA and transfer RNA. 94 Created by Julia Hsu Levy – Version 1.5 * * * Translation: Protein Synthesis * * * Translation is the process of ribosomes forming peptide bonds between amino acids, using mRNA as the template. The result is the production of proteins. Ribosomes are created from rRNA, ribosomal RNA. Two strands of rRNA fold to make the large and the small subunit of the ribosomal complex. Prokaryote: 70S = 30S + 50S Eukaryote: 80S = 40S + 60S Codon: sequence of three RNA nucleotides that correspond with an anticodon of a transfer RNA molecule, carrying an amino acid 1. initiation: Transfer RNA activation: tRNA strands are “charged” with a corresponding amino acid. GTP dephosphorylates to GDP, supply the necessary “energy” to fuel this reaction. The small and large ribosomal subunits are separated by initiation factors (proteins). f-met: formylmethionyl-tRNA binds to the smaller ribosomal subunit and together bind to the initiator codon AUG. The larger subunit binds to the initiator complex, and translation begins. The larger subunit has two “pockets” or regions named for their functions: P 95 Created by Julia Hsu Levy – Version 1.5 (transpeptidation) and A (acceptor) sites. The f-met-tRNA is positioned in the Psite. 2. Elongation: Elongation factors locate the appropriate t-RNA with the matching anticodon of the codon near the Asite. An enzyme called peptidyl transferase to the A-site tRNA transfers the amino acid of the t-RNA in the P site. Peptide bonds link the two amino acids. Translocation: The ribosome shifts such that the A-site tRNA is now in the P-site. The process repeats, and the amino acid chain grows. The process stops when the ribosome encounters a codon that indicates “stop.” 3. Termination: A water molecule dissociates the polypeptide chain from the t-RNA. 96 Created by Julia Hsu Levy – Version 1.5 97 Created by Julia Hsu Levy – Version 1.5 * * * Operons * * * Central dogma: Genes (DNA) code for the production of amino acids, which fold into proteins and are expressed as the phenotype of an individual. Genes also code for the production of protein enzymes. Assuming no mutations, how is gene expression regulated? 1. Feedback inhibition: If too much gene is expressed and there is ample supply of the necessary protein, the product of the metabolic pathway will inhibit its own production. Example: The cell produces cyclin at the G-checkpoints. If cyclin levels are high, the cell will initiate the breakdown of its cyclin to slow down (regulate) the release of MPF. Example: All bacteria require tryptophan (an amino acid –trp) to survive. If the bacteria does not find trp in the environment, it must synthesize its own trp. If too much trp has been made, to conserve energy, the high [trp] signals the pathway to stop. 2. Enzymatic inhibition: Cells can adjust for its own enzyme catalytic levels by introducing allosteric or non-allosteric inhibition. How organisms control gene expression: Operons: transcription units that can consist of multiple genes (polycistronic) or a single gene (monocistronic) Polycistronic operons (bacteria) are made up of a promoter region, an operator, and the genes, which they control. They control genes by producing the necessary enzymes that allow or prevent transcription. Trp operon: a pathway for regulating tryptophan production The operator is always set to “go.”. It allows for the RNA polymerase holoenzyme to precede transcription. The operator is always on. To turn the operator off, the operon must have a repressor protein. The repressor protein is made with a regulatory gene 98 Created by Julia Hsu Levy – Version 1.5 that is found upstream of the operator. Tryptophan acts as a corepressor to turn on the repressor protein. The trp operon is repressible Lac operon: a pathway for breaking down lactose (disaccharide sugar) into galactose and glucose for energy The operator is always set to “stop.” It does not allow for the RNA polymerase holoenzyme to proceed transcription. A repressor is always “on” the operator. An inducer molecule binds to the repressor to inactivate the repressor. The inducer is allolactose. In the presence of lactose, allolactose binds to the operator to turn it on. The lac operon is inducible, which means that it must be induced to begin transcription. Positive regulation: cAMP (cyclic AMP) builds up in concentration when glucose is absent. CRP (cAMP regulatory protein) binds to the allosteric site of CRP. This activates transcription. The CRP+ cAMP binds to the promoter region of the DNA, this helps the RNA polymerase to bind to the promoter. 99 Created by Julia Hsu Levy – Version 1.5 * * * Mutation * * * Mutations are any chances in the genetic makeup of an organism. Mutations do not lead to disorders in every situation. Point mutations: changes to any one nucleotide Substitutions – Polymerase may add one nucleotide in place of the correct one. -Due to redundancy (wobble), the mutation may not lead to a change in the amino acid sequence. These are called silent mutations. Insertions – frameshift mutation; Because of the addition of a nucleotide, the ribosomes will read a different sequence of codons than intended. Deletions – frameshift mutation; Because of the subtraction of a nucleotide, the ribosomes will read a different sequence of codons than intended. How do mutations occur? 1. Mistakes during DNA replication 2. Mistakes during recombination (crossing-over) 3. Mistakes during transcription 4. Mistakes during translation 5. Mutations: chemical or physical substances that can cause DNA mutations X-rays UV thymine dimers High energy radiation Base analogues – chemicals with similar chemical structure as DNA nucleotides 100 Created by Julia Hsu Levy – Version 1.5 Ames test: simple testing method developed by Bruce Ames to test for the mutagenic activity of certain chemicals How are DNA mutations corrected? Excision repair Removal of lesions Postreplication repair Recombinant repair 101 Created by Julia Hsu Levy – Version 1.5 Chapter 20: Biotechnology Somatic nuclear transfer (nuclear transplantation): The nucleus is removed from a differentiated somatic cell and transplanted into an empty egg cell. UV light is used to destroy the nucleus in the egg cell. The transplanted egg cell is forced (with additions of hormones, etc) to undergo embryonic development. Dolly and Polly the sheep were both cloned with this technology. Recombinant plasmids: altering the genetic makeup of plasmids (circular pieces of bacterial DNA) 102 Created by Julia Hsu Levy – Version 1.5 Transformation: inserting pieces of bacterial DNA into a bacterium, altering its genetic makeup Phage libraries Plasmid libraries: cDNA libraries: complementary DNA libraries 103 Created by Julia Hsu Levy – Version 1.5 Polymerase chain reaction (PCR): technique for making DNA libraries (multiple copies) using polymerase enzyme and thermal cycling Gel electrophoresis in RFLP technology: RFLP: Restriction fragment length polymorphism Most of the genome consists of non-coding (“junk”) DNA. These junk sequences are heritable in the same ways that coding DNA (genes) are. It is fair to say that two related organisms should share similar sequences of junk DNA. Restriction enzymes were isolated from bacteria, which had them as defense against viruses. Viruses cleave their genetic material into the bacterial genome, and restriction enzymes are used to cut out the viral DNA. Restriction enzymes only recognize specific DNA sequences called palindromes. Ex: 3’ – AGGCCT – 5’ 5’ – TCCGGA – 3’ 104 Created by Julia Hsu Levy – Version 1.5 The sequences read the same in a 3’ to 5’ direction. The enzyme would break the phosphodiester bonds between the A and G, making a zig-zag cut, leaving “sticky ends.” If the palindrome sequence is CCGG the cut would be made between C and G. GGCC The resulting cut would leave “blunt ends.” If two pieces of DNA are treated to the same restriction enzyme, chances are that they would have palindrome sequences in nearly the same places in their non-coding DNA if the DNA are related. The result of treating with restriction enzymes is the creation of DNA fragments of various lengths (RFLPs). The DNA fragments of each sample are loaded into the wells of an agarose gel. The gel is placed into an electrophoresis chamber with a running buffer solution, and plugged into an electric source to generate a voltage differential gradient. The electric charge separates the DNA bands in a process called gel electrophoresis. RFLP analysis: 105 Created by Julia Hsu Levy – Version 1.5 Sanger method: Southern blotting: What are ways in which these technologies can help human societies? (gene therapy?) 106