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AP Biology
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Genetics Part 2B
Regulation of metabolic pathways Operon: cluster of related genes with on/off switch
Three Parts:
1. Promoter – where RNA polymerase attaches
2. Operator – “on/off”, controls access of RNA poly
3. Genes – code for related enzymes in a pathway
Bacterial control of gene expression
Regulatory gene: produces repressor protein that binds to operator to block RNA poly
Repressible Operon (ON OFF) Repressible Operon
Normally ON
Anabolic (build organic molecules)
Organic molecule product acts as corepressor binds to repressor to activate it
Operon is turned OFF
Eg. trp operon
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trp operon
Inducible Operon
Normally OFF
Catabolic (break down food for energy)
Repressor is active inducer binds to and inactivates repressor
Operon is turned ON
Eg. lac operon
lac operon
• Typical human cell: only 20% of genes expressed at any given time
• Different cell types (with identical genomes) turn on different genes to carry out specific functions
• Differences between cell types is due to differential gene expression
Eukaryotic gene expression
regulated at different stages
Chromatin Structure:
• Tightly bound DNA less accessible for transcription
• DNA methylation: methyl groups added to DNA; tightly packed; transcription
• Histone acetylation: acetyl groups added to histones; loosened; transcription
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Transcription Initiation: • Control elements bind
transcription factors • Enhances gene expression
Transcription Initiation Complex
Enhancer regions bound to promoter region by activators
Regulation of mRNA:
• micro RNAs (miRNAs) and small interfering RNAs (siRNAs) can bind to mRNA and degrade it or block translation
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1. Cell Division: large # identical cells through mitosis
2. Cell Differentiation: cells become specialized in structure & function
3. Morphogenesis: “creation of form” – organism’s shape
Embryonic Development: Zygote Organism
Determination: irreversible series of events that lead to cell differentiation
• Cytoplasmic determinants: maternal substances in egg distributed unevenly in early cells of embryo
• Induction: cells triggered to differentiate
• Cell-Cell Signals: molecules produced by one cell influences neighboring cells – Eg. Growth factors
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Pattern formation: setting up the body plan (head, tail, L/R, back, front)
Morphogens: substances that establish an embryo’s axes
Homeotic genes: master control genes that control pattern formation (eg. Hox genes)
1. Proto-oncogene = stimulates cell division
2. Tumor-suppressor gene = inhibits cell division
• Mutations in these genes can lead to cancer
Control of Cell Cycle: Proto-oncogene Oncogene
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Genes involved in cancer:
• Ras gene: stimulates cell cycle (proto-oncogene)
– Mutations of ras occurs in 30% of cancers
• p53 gene: tumor-suppresor gene
– Functions: halt cell cycle for DNA repair, turn on DNA repair, activate apoptosis (cell death)
– Mutations of p53 in 50+% of cancers
• Cancer results when mutations accumulate (5-7 changes in DNA)
• Active oncogenes + loss of tumor-suppressor genes
• The longer we live, the more likely that cancer might develop
Bacteria vs. Viruses
Bacteria Virus
• Prokaryotic cell
• Most are free-living (some
parasitic)
• Relatively large size
• Antibiotics used to kill
bacteria
• Not a living cell (genes
packaged in protein shell)
• Intracellular parasite
• 1/1000 size of bacteria
• Vaccines used to prevent
viral infection
• Antiviral treatment
Viruses
• Very small (<ribosomes)
• Components = nucleic acid + capsid
– Nucleic acid: DNA or RNA (double or single-stranded)
– Capsid: protein shell
– Some viruses also have viral envelopes that surround capsid
• Limited host range (eg. human cold virus infects upper respiratory tract)
• Reproduce within host cells
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Simplified viral replicative cycle Bacteriophage
• Virus that infects bacterial cells
Lytic Cycle of T4
Phage Bacteriophage Reproduction
• Lytic Cycle:
– Use host machinery to make copies of virus
– Death of host cell by rupturing it (lysis)
– Virulent phages replicate by this method
• Lysogenic Cycle:
– Phage DNA incorporated into host DNA and replicated along with it
– Phage DNA = prophage
• Temperate Phage: uses both methods of replication
Lytic Cycle vs. Lysogenic Cycle
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Animal viruses have a membranous envelope
• Host membrane forms
around exiting virus
• Difficult for host
immune system to
detect virus
Retrovirus
• RNA virus that uses reverse transcriptase (RNA
DNA)
• Newly made viral DNA inserted into chromosome
of host
• Host transcribes viral DNA (= provirus) to make
new virus parts
• Example: HIV (Human Immunodeficiency Virus)
HIV =
Retrovirus
HIV
◦ Infects white blood cells
◦ HIV+: provirus (DNA inserted)
◦ AIDS: active viral reproduction
Vaccines
• Weakened virus or part of pathogen that
triggers immune system response
Emerging viruses = mutation of existing
viruses
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Viroids
• Small, circular RNA
molecules that infect plants
• Cause errors in regulatory
systems that control plant
growth
• Eg. coconut palms in
Philippines
Prions
• Misfolded, infectious proteins that cause
misfolding of normal proteins
• Eg. mad cow disease (BSE),Creutzfeldt-Jakob
disease (humans), scrapie (sheep)
Diseases caused by prions
• Prions act slowly – incubation period of at least 10
years before symptoms develop
• Prions are virtually indestructible (cannot be
denatured by heating)
• No known cure for prion diseases
Tools of Genetic Engineering
Restriction enzymes (restriction endonucleases): used to cut strands of DNA at specific locations (restriction sites)
Restriction Fragments: have at least 1 sticky end (single-stranded end)
DNA ligase: joins DNA fragments
Cloning vector: carries the DNA sequence to be cloned (eg. bacterial plasmid)
Using a restriction
enzyme (RE) and
DNA ligase to
make recombinant
DNA
Gene Cloning
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Applications of Gene Cloning
PCR (Polymerase Chain
Reaction): amplify (copy)
piece of DNA without use
of cells
Gel Electrophoresis: used to separate DNA molecules on
basis of size and charge using an electrical current (DNA
+ pole)
Gel Electrophoresis: used to separate DNA molecules on
basis of size and charge using an electrical current (DNA
+ pole)
Cloning Organisms
• Nuclear transplantation: nucleus of egg is removed and replaced with nucleus of body cell
Nuclear Transplantation
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Problems with Reproductive Cloning
• Cloned embryos exhibited various defects
• DNA of fully differentiated cell have epigenetic changes
Stem Cells
• Stem cells: can reproduce itself indefinitely and
produce other specialized cells
– Zygote = totipotent (any type of cell)
– Embryonic stem cells = pluripotent (many cell types)
– Adult stem cells = multipotent (a few cell types) or
induced pluripotent, iPS (forced to be pluripotent)
Embryonic
vs. Adult
stem cells
Applications of DNA Technology
1. Diagnosis of disease – identify alleles, viral DNA
2. Gene therapy – alter afflicted genes
3. Production of pharmaceuticals
4. Forensic applications – DNA profiling
5. Environmental cleanup – use microorganisms
6. Agricultural applications - GMOs
Gene therapy using a retroviral vector “Pharm” animal: produce human protein
secreted in milk for medical use
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DNA Fingerprinting RFLPs (“rif-lips”)
• Restriction Fragment Length Polymorphism
• Cut DNA with different restriction enzymes
• Each person has different #s of DNA fragments created
• Analyze DNA samples on a gel for disease diagnosis
• Outdated method of DNA profiling (required a quarter-sized sample of blood)
RFLPs – Disease Diagnosis
STR Analysis
• STR = Short Tandem Repeats
• Non-coding DNA has regions with
sequences (2-5 base length) that are repeated
• Each person has different # of repeats at
different locations (loci)
• Current method of DNA fingerprinting used
– only need 20 cells for analysis
STR Analysis STR Analysis
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Human DNA
• 3 billion base pairs
• ~20,000 genes
• Only 1.5% codes for proteins (or RNA)
• Repetitive DNA: sequences present in multiple copies
Transposable Elements
Make up 75% of repetitive DNA
Can be moved from one location to another in genome
Discovered by Barbara McClintock – corn breeding experiments
2 Types:
Transposons
Retrotransposons
Transposons
Moves within genome via DNA intermediate
“cut & paste” or “copy & paste” mechanisms
Requires enzyme transposase
Retrotransposons
Move by means of RNA intermediate
Leaves copy at original site
Involves enzyme reverse transcriptase
Multigene Families
• Collections of 2 or more identical or very similar genes
• Eg. hemoglobin: -globin and -globin gene families
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Transpositions chromosomal rearrangements Transposable elements contribute to evolution
• Promote recombination, disrupt genes or control elements, & carry genes to new locations
• May be harmful or lethal, but can also have small beneficial effects
• Provides raw material for natural selection
Evolutionary Development (Evo-devo)
• Compares developmental processes to understand how changes can lead to evolution of organisms
Homeotic genes: master regulatory genes
• Control placement and spatial organization of body parts
Homeobox: widely conserved 180-nucleotide sequence within homeotic (Hox) genes
• Found in many groups (fungi, animals, plants)
• Hints at relatedness between all life forms
Conservation of homeotic genes
Genetic Diversity in Prokaryotes
Factors:
1.Rapid reproduction (binary fission)
2.Mutations – errors in replication
3.Genetic recombination
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Genetic Recombination in Bacteria
1. Transformation: uptake of foreign DNA from surroundings
2. Transduction: viruses transfer genes between prokaryotes
3. Conjugation: DNA transferred from one to another
Transformation • Uptake of foreign DNA from surroundings
• Observed by Griffith (bacteria & mice)
Plasmids
Small ring of DNA that carries a few genes
Replicates separately from bacterial chromosome
Can carry genes for antibiotic resistance
Used frequently in genetic engineering for gene cloning
AP Bio Lab 6A - Transformation Using plasmids and bacteria in genetic engineering
Transduction
• Viruses (bacteriophages) carry bacterial genes from one host cell to another
• Recombine DNA of donor and recipient cell
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Conjugation
• One cell donates DNA to another
• Donors cell extends a sex pilus (“mating bridge”) through which DNA is transferred
• Requires the presence of a piece of DNA called the F factor to produce the pilus