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Molecular Biology Introduction – Definitions – History – Central Dogma Molecular Biology Definitions – – – – – Chromosomes DNA Gene Genotype Phenotype Molecular Biology Chromosomes – The structure in cells that carries hereditary information – Composed of DNA and protein – Prokaryotic - circular – Eukaryotic - linear Molecular Biology DNA – – – – Nitrogenous base (4 bases - A,T,G & C Deoxyribose sugar Phosphate Nitrogenous bases are paired » AT » GC – Double helix structure Molecular Biology Genes – Segments of DNA – Functional or regulatory – Mutability and variation Molecular Biology Genotype – The genetic make-up of an organism; the information that codes for all the characteristics of an organism Phenotype – The expression or physical manifestation of a gene; how it appears Molecular Biology Molecular biology seeks to understand the molecular or chemical basis of genetics History of molecular biology is a melding of biochemistry, especially nucleic acid biochemistry and genetics Molecular Biology Biochemistry – – – – Meischer Avery & MacLeod Hershey & Chase Watson & Crick Genetics – – – – – – – Mendel Sutton Morgan Griffith Delbruck Beadle & Tatum Tatum & Lederberg Molecular Biology - Genetics Mendel (1865) – – – – – Fluid vs. particulate inheritance Studied pure breeding pea plants Law of Segregation Law of Independent Assortment Rediscovered by de Vries & others Molecular Biology Molecular Biology Molecular Biology - Genetics Cross of pure breeding purple flowers with pure breeding white flowers produces all purple plants with genotype Pp; crossing Pp plants produces following distribution: P p P PP purple Pp purple p Pp purple pp white Molecular Biology - Genetics Walter Sutton (1902) – Studied meiosis in grasshoppers (insects have large readily observable chomosomes) – Observed that chromosomes behave in manner similar to segregation of hereditary material – Found that chromosomes occur in morphologically similar pairs – Pairs separate during meiosis Molecular Biology - Genetics Molecular Biology - Genetics Morgan – Developed modern science of genetics – Used fruit flies because they had a shorter generation time than peas – Discovered sex-linkage – Students developed techniques of mapping genes on chromosomes Molecular Biology - Genetics Griffith – discovered transformation in 1927 – is a means of genetic transfer in microorganisms – a process by which a nonpathogenic strain is transformed into a pathogenic strain Molecular Biology - Genetics Molecular Biology - Genetics Delbruck – developed quantitative methods for analysis of bacteriophage; viruses of bacteria – organized course to teach biologists methods at Cold Spring Harbor resulting in a large number of biologists trained in molecular techniques Molecular Biology - Genetics Beadle & Tatum – developed Neurospora as an experimental organism – established one gene one enzyme hypothesis – generation time is even shorter with Neurospora Molecular Biology - Genetics Molecular Biology - Genetics Molecular Biology - Genetics Tatum & Lederburg – discovered conjugation in bacteria Molecular Biology - Biochemistry Meischer (1869) – Austrian doctor – isolated a substance called “nuclein” from the nuclei of cells obtained from the pus of surgical bandages – found to contain nitrogenous chemicals, sugar and phosphate Molecular Biology - Biochemistry Avery & MacLeod (1944) – isolated Griffith’s transforming factor to a high degree of purity – characterized transforming factor using highly purified enzymes – found transforming factor to be DNA Molecular Biology Substance Enzyme Transform Capsule Carbohydrase Yes Protein Protease Yes RNA RNAse Yes DNA DNAse No Molecular Biology - Biochemistry Hershey & Chase (1952) – used newly developed radioisotopes » 35S for protein » 32P for nucleic acid – labeled bacteriophage (a virus of bacteria) – found 32P went into cells but 35S did not implying that nucleic acid transfer information to cell for new bacteriophages Molecular Biology - Biochemistry Watson & Crick (1953) – used X-ray crystallography to study structure of DNA – by combining chemical data and X-ray data were able to construct a model of DNA – structure inferred function leading to Central Dogma Molecular Biology Central Dogma – – – – – DNA Structure Genetic Code Replication Transcription Translation Molecular Biology DNA Structure – – – – – Sugars Bases Phosphates Double Helix Anti-parallel Molecular Biology Genetic Code – 4 bases / 20 amino acids – codons – punctuation Molecular Biology Universal Genetic Code Molecular Biology Central Dogma states a hypothesis regarding information flow in cell Replication - the copying of DNA or information for next generation Transcription - the copying of information for use by the cell Translation - the conversion of information into useful products enzymes Molecular Biology Replication Transcription DNA DNA Polymerase Translation mRNA RNA Polymerase mRNA tRNA Ribosomes Protein Molecular Biology Replication – – – – – – replication is semi-conservative replication occurs at replication fork replication is discontinuous process uses DNA polymerase Uses RNA polymerase requires a primer with free 3’-hydroxyl Molecular Biology Molecular Biology The Meselson Stahl Experiment Molecular Biology Molecular Biology Molecular Biology Continuous/Discontinuous DNA Synthesis Molecular Biology Transcription – – – – RNA polymerase promoters produces messenger RNA (mRNA) requires NO primer Molecular Biology Molecular Biology Differences between prokaryotic and eukaryotic mRNA Prokaryotic – often polygenic – turns over quickly – translated almost immediately Eukaryotic post transcription modification – Heterogenous introns & exons - excision of introns – mRNA stability – days to weeks – Addition of 5’ cap and 3’ polyadenylation Molecular Biology Translation – – – – – tRNA amino acid synthase ribosomes initiation termination Molecular Biology Molecular Biology DNA Sequence Analysis – DNA coding (sense) » 5’CCG ATG AAT GTC GAG CTA TCC TAC 3’ – DNA non-coding (nonsense)(template) » 3’GGC TAC TTA CAG CTC GAT AGG ATG 5’ – mRNA » 5’CCG AUG AAU GUC GAG CUA UCC UAC 3’