Ap Biology – Modern Genetics
DNA is the template (3’ to 5’) ! 3. RNA Polymerase – Binds to promoters, separates strands of DNA and ! hook together RNA nucleotides (only add to 3’ end) ! a. TATA Box – Where RNA polymerase H binds (eukaryotes) ! ! b. Transcription Factors – Aid polymerase in locating promoters ! ! such as the TATA Box ! ! c. Reads in sets of 3 called codons. ! 4. Termination site on DNA indicates end in prokaryotes ! 5. Eukaryotes – Pre-mRNA separates from DNA at the polyadenylation ! signal, but RNA polymerase continues to transcribe until it falls off the ! DNA. C. Pre-mRNA processing in eukaryotes ! 1. Heterogenous nuclear RNA (hnRNA) contains introns and exons. ! ! a. Introns not used (stay IN nucleus) ! ! b. Exons used (EXIT nucleus) ! 2.
Small nuclear ribonucleic proteins (snRNP’s/snurps) help excise introns ! and fuse exons. ! 3. Splicosomes – Snurps and proteins working together ! 4. Poly A tail is added to 3’ end which a 5’ cap is added to the 5’ end. ! ! a. Prevent damage to mRNA as it leaves nucleus ! ! b. Helps mRNA leave nucleus ! ! c. Helps mRNA attach to ribosome. D. rRNA – Formed in nucleoli, makes up part of ribosome. ! 1. Ribosomes – Binding site for rRNA. E. Translation ! 1. Initiation – Brings mRNA and tRNA together by means of factors (start ! codon is AUG) ! 2. Elongation – Amino acids are added with aide of elongation factors. ! ! a.
Codon Recognition – mRNA codon on A site forms H bonds with ! ! anticodon of incoming tRNA. ! ! b. Aminoacyl tRNA synthetase matches amino acid with correct ! ! tRNA ! ! c. Peptidyl transferase catalyzes the formation of a peptide bond ! ! between polypeptide at P site and amino acids at A site. ! ! d. Wobble Effect – Exact base pairing is not always necessary. ! 3. Translocation ! ! a. A site on ribosome holds tRNA carrying the next amino acid to be ! ! added. ! ! b. P site on ribosome holds tRNA carrying the growing polypeptide ! ! chain. ! ! c. E site is where it exits. F. Termination – Indicated by termination codons (UAA, UAG, UGA) ! . Release Factor – Protein which binds to termination codon on A site ! causing hydrolysis reaction and freeing the polypeptide. ! 2. Posttranslational modi? cations now take place – Protein goes into ! secondary, and tertiary structure. AP Bio – Modern Genetics IV. Mutations ! A. Point Mutations – Chemical changes in one or a few nucleotides. ! ! 1. Base Pair Substitutions – Replacement of one nucleotide and its partner ! ! from a complementary DNA strand with another pair of nucleotides. ! ! ! a. Missense Mutations – Altered codons still code from amino acids ! ! ! but don’t always make sense. ! ! b. Nonsense Mutations – Change amino acids codon to a stop ! ! ! signal (most result in nonfunctional codons) ! ! 2. Insertions and Deletions – Additions or losses of one or more nucleotide ! ! pairs in a gene. ! ! ! a. Frameshift Mutation – Alters reading from (usually nonfunctional) ! B. Mutagenesis – Creation of mutations ! ! 1. Hermann Muller discovered that genetic changes occurred when fruit ! ! ? ies were exposed to X-Rays. ! ! 2. Spontaneous mutations result from errors in DNA replication, repair, or ! ! recombination. ! ! 3. Mutagens – Physical and chemical agent which cause mutations.
Chapter 18 – The Genetics of Viruses and Bacteria I. Viruses ! A. Wendell Stanley (American-1935) crystallized particle, now called tobacco ! mosaic virus (TMV) ! B. Genomes – May be double or single stranded DNA or RNA. ! C. Capsid – Protein shell enclosing genome, built from capsomere ! D. Phages – Virus which infects bacteria. ! E. Lack enzymes for metabolism and have no ribosomes, nonliving ! F. Vaccine – Harmless variation which stimulates immune response (? rst made ! from cowpox) ! G. Viroids – Tiny molecules of naked RNA, upset metabolism of plants ! H. Prions – Infectious proteins ! ! 1.
Often cause brain diseases like mad cow and scrapie in sheep ! ! 2. Not killed by heat, long incubation period II. Sources of New Viruses ! A. Mutations of current viruses ! B. Spread between species typically due to new exposure. ! C. Spread within species between a small population and the rest of the world. III. Replication and Infection ! A. Obligate intracellular parasites ! B. Basic Infection ! ! 1. Some recognize host by a “lock and key” ? t, and only infect certain cells ! ! of the body. ! ! 2. Genome Enters Cell – Method varies (leaving ghost behind-empty ! ! capsid) AP Bio – Modern Genetics ! ! ! ! ! ! ! ! ! ! ! ! ! 3. Genome reprograms cell to copy viral genes and manufacture capsid ! proteins. ! ! a. DNA viruses use DNA polymerases of host cell ! ! b. RNA viruses contain their own enzymes to initiate replication. ! 4. Self Assembly – Assembly of capsids and nucleic acid molecules into ! viruses. ! 5. Viruses leave host cell C. Bacteriophages – Virus which infect bacteria ! 1. Lytic Cycle – Reproductive cycle which ends with death of host cell ! 2. Lysogenic Cycle – Reproduces viral genome without destroying host ! ! a. Viral DNA is incorporated into bacterial DNA (prophage) ! b. Prophage is replicated every time bacteria devices ! ! c. Eventually goes into lytic cycle. IV. Animal Viruses ! A. Viruses with Envelopes ! ! 1. Envelopes fuse with host membrane, inserting information ! ! 2. Enzymes remove capsid, genome replicates ! ! 3. Viral offspring are wrapped in a membrane and released ! ! 4. Provirus – Some viruses become integrated into cell’s genome (infection ! ! tends to reoccur throughout life, HIV, herpes) ! B. RNA Viruses ! ! 1. Retroviruses – Most complicated life cycle (HIV) ! ! ! a. Reverse Transcriptase – Transcribes DNA from RNA template ! ! ! b.
Newly formed DNA integrates as a provirus into a chromosome ! ! ! within the nucleus of the host cell. ! C. Viral Diseases ! ! 1. Vaccine – Harmless variants or derivatives of pathogenic microbes ! ! which stimulate an immune response. V. Plant Viruses ! A. Horizontal Transmission – Plant infected from external source ! B. Vertical Transmission – Plant inherits infection from parent VI. Bacteria ! A. Single stranded, circular DNA, found in nucleoid region ! B. Plasmids – Smaller circles of DNA, used in genetic engineering ! C. Replication of chromosome is followed by binary ? ssion (copying of DNA ! occurs in both directions) VII.
Genetic Recombination of Bacteria ! A. Transformation – Alteration of a bacterial cell’s genotype by the uptake of ! naked, foreign DNA from environment. ! B. Transduction – Transfer of bacteria by means of a virus ! C. Conjugation – Direct transfer of genetic material between two bacterial cells ! ! 1. Donor is “male” (requires F plasmid, F+), receiver is “female” (F-) AP Bio – Modern Genetics ! ! ! ! ! ! ! ! ! ! ! ! ! ! ! ! 2. Plasmid – Not required for survival or reproduction, but can be ! advantageous in stressful environments. ! ! a. Episomes – Can replicate as a free molecule in cytoplasm or as ! part of the main bacterial chromosome. ! 3. R Plasmids cause bacteria to be resistant to antibiotics D. Transposons – Pieces of DNA which can move from one location to another in a cell’s genome (jumping genes) discovered by Barbara McClintock. ! 1. Conservative Transposition – Genes are not replicated, number of ! copies is conserved (cut and paste) ! 2. Replicative Transposition – Transposon replicates at original site and a ! copy inserts at another location (copy and paste) E. Insertion Sequences – Simplest transposons, consist of only the DNA necessary for act of transposition itself.
F. Barbara McClintock (1940’s-50’s) worked with Indian corn, based on observations, she believed there were mobile elements. VII. Operons – Unit of genetic function common in bacteria and phages and consisting of regulated clusters of genes with related function. ! A. Jacob and Monod – Discovered operon model in 1961 ! B. Operon is made of an operator, promoter and the genes they control ! C. Operator – Switch which gets turned on ! D. Oppressor – Segment of DNA, functions as an on/off switch for an operon ! E. Repressor – Protein which turns off the operon (operator speci? c) !
F. Corepressor – Metabolite that cooperates with a repressor protein to switch an ! operon off ! G. E coli Operons ! ! 1. Lac Operon ! ! ! a. Regulatory Gene —> Repressor —> Binds to Operator ! ! ! b. RNA Polymerase cannot transcribe structural genes needed to ! ! ! make enzymes break down lactose ! ! ! c. When lactose is present, some of the lactose attaches to the ! ! ! repressor and shuts it down allowing RNA polymerase to transcribe ! ! ! (example of an inducible operon – Usually off but can be induced. ) ! ! 2. Trp Operon (Tryptophan is an amino acid) ! ! ! a.
Regulatory Gene —> Inactive repressor that doesn’t attach to the ! ! ! operator so RNA polymerase can transcribe. ! ! ! b. Increased levels of trp combine with repressor to make it active ! ! ! and the RNA polymerase can’t transcribe (example of corepressor) ! ! ! c. Repressible Operon – Transcription usually on but can be ! ! ! repressed Chapter 19 – Eukaryotic Genomes: Organization, Regulation, and Evolution I. Key Terms ! A. Cellular Differentiation – Process of cells becoming specialized and therefore ! only parts of genes expressed. AP Bio – Modern Genetics ! ! ! ! ! ! ! ! ! B.
Histones – Positively charged proteins which bind to negative DNA and “package it together” C. Nucleosome – Basic unit of DNA packaging in eukaryotes, consisting of DNA around a protein core (2 copies of each of the 4 types of histones) D. Multigene Families – Collections of identical or similar genes (hemoglobin alpha and beta are expressed at different times) E. Pseudogenes – Similar to functioning genes but are lacking some essential part. F. Heterochromatin, euchromatin, telomere. II. Gene Control – Eukaryotes ! A. Transcriptional Control – Factor which tell where to start and stop ! B. Posttranscriptional – Splicing, cap and tail !
C. mRNA Degradation ! ! 1. Prokaryotic mRNA are usually broken down within minutes of use ! ! 2. Eukaryotic mRNA can live hours to weeks before being broken down by ! ! enzymes. ! D. Chromosomes Puffs – In insects, regions of intense transcription, chemicals ! signal when they should form. ! E. Steroids in Humans ! ! 1. Steroid diffuses through cell (lipid soluble) into cytoplasm and binds to ! ! receptor. ! ! 2. Binding to an enhancer then occurs and activates transcription. ! ! 3. Steroid acts as a signal to turn on speci? c gene. ! F. Hormones bind to outside of cell (not lipid soluble) and operate with the help of ! receptor protein on the outside of the cell. ! G. Gene Ampli? cation – Selective replication of certain genes. ! H. Immunoglobulin – Proteins which speci? cally recognize and help ? ght viruses, ! bacteria and invaders. ! ! 1. Made by B lymphocytes ! ! 2. Antibody variation results from different combinations of variable and ! ! constant regions of immunoglobulin polypeptides. ! I. DNA Methylation III. Abnormal Expression of Genes ! A. Oncogenes – Cancer causing genes found in RNA viruses ! B. Proto-Oncogenes – Found in human cells, normal genes which may become ! oncogenes. ! ! 1. Increased activity may cause change ! 2. Gene Ampli? cation ! ! 3-5. Translocation, Gene Transposition, and Point Mutation ! C. Tumor Suppressor Genes – Normally prevent cell division and may produce ! proteins which repair damaged DNA. ! D. The more mutations throughout life, the greater the chance of cancer. AP Bio – Modern Genetics Chapter 20 – DNA Technology I. Gene Manipulation ! A. Restriction Enzymes – Bacterial enzymes which cut up foreign DNA forming ! sticky ends. ! ! 1. Sticky ends can be used to join DNA pieces originating from different ! ! sources. ! ! 2. Restriction fragments can be separated by gel electrophoresis ! ! ! a.
Brief electric pulse is applied to a solution of DNA to separate ! ! ! DNA. ! ! ! b. DNA is negative and moves towards positive end ! ! ! c. Can be used to compare DNA of different species. ! B. Gene Cloning in a Plasmid ! ! 1. Isolate bacterial plasmid and eukaryotic DNA with gene desired ! ! 2. Both are treated with same restriction enzyme ! ! ! a. Plasmid DNA – Cut at restriction site ! ! ! b. Eukaryotic DNA – Generates thousands of fragments ! ! 3. Two types of DNA are mixed – Sticky ends of each pair up ! ! 4. DNA ligase joins molecules (called recombinant DNA) ! ! 5. Recombinant DNA is introduced into a bacterial cell ! 6. Gene Cloning – Production of multiple copies of desired gene ! C. Inserting DNA into Cells ! ! 1. Cloning Vectors – Carriers for moving recombinant DNA (plasmids, ! ! phages-transformation) ! D. Sources of Genes for Cloning ! ! 1. DNA isolated directly from organism ! ! 2. Complementary DNA (cDNA) made in a lab from mRNA templates. ! E. Probes – Nucleic molecules which will hydrogen bond speci? cally to a desired ! gene, location traced by labeling ! F. Sanger Method of Sequencing ! ! 1. Synthesizing in vitro DNA strands complementary to one of the strands ! ! being sequenced. ! G. Amplifying DNA ! ! 1.
Polymerase chain reaction (PCR) technique by which any piece of DNA ! ! can be quickly ampli? ed in vitro ! ! ! a. DNA is incubated with DNA polymerase and primers. II. Applications ! A. Human Genome Project – Mapping of entire human genome ! B. Analyzing genomes of other species ! C. Human Gene Therapy – Correct Disorders ! D. Vaccines ! E. Mammalian Hormones and Proteins – Growth hormones, clotting factor, insulin ! F. Treating Farm Animal with vaccines, antibodies and growth hormones ! G. Transgenic Organisms – Contain genes from another species ! H. Manipulating Plant Genes – Herbicide resistance, better foods