1. Lecture 1 & 2: DNA, Genes, Chromatin Learning Objectives Understand the structure of DNA, including the structure of the bases, nucleosides, nucleotides, and the structure of the DNA double helix Understand the significance of the specificity of base pairing and the complementarily of the DNA strands Understand the nature of the forces contributing to the stability of the DNA double helix Understand the process of DNA denaturation and the relationship between melting temperature and the base composition of DNA Understand what repetitive sequences are and how they are arranged in the human genome Understand the mechanism by which Alu sequences have affected the LDL receptor gene Understand basic gene structure and the basic characteristics of human nuclear and mitochondrial DNA Understand basic chromosome structure and how DNA is packaged into chromosomes Understand the significance of telomeres to aging and cancer and the use of telomerase in cancer diagnosis 1

2. GENETIC DOGMA Genetic diseases occur because of mutations in DNA. Many of these mutations affect the repair of other mutations that occur during DNA replication or at other times, which in turn affect the flow of genetic information from DNA to RNA (transcription and processing) and from RNA to protein synthesis (translation). Many of these mutations also affect the structures of the resulting proteins, affecting their functions. 2

3. 3 THE FLOW OF GENETIC INFORMATION DNARNAPROTEIN DNA 1 23 1. REPLICATION(DNA SYNTHESIS) 2. TRANSCRIPTION(RNA SYNTHESIS) 3. TRANSLATION(PROTEIN SYNTHESIS)

4. 4 Evidences that DNA is the genetic material, DNA Structure and Chemistry a). Evidence that DNA is the genetic information i). DNA transformation – know this term ii). Transgenic experiments – know this process iii). Mutation alters phenotype – be able to define genotype and phenotype b). Structure of DNA i). Structure of the bases, nucleosides, and nucleotides ii). Structure of the DNA double helix iii). Complementarity of the DNA strands c). Chemistry of DNA i). Forces contributing to the stability of the double helix ii). Denaturation of DNA

5. Evidences that DNA is the genetic material I.a. DNA transformation Experiment: Injecting mice with both: a heat-killed virulent strain + a non-heated, non-virulent strain Streptococcus. Result and interpretation: Something (DNA) from the heat-killed virulent strain of Streptococcus was able to alter the (still viable) non- virulent strain, converting some of the cells to virulent bacteria and killing the host. 5

6. I.b. DNA transformation. DNA is the carrier of genetic information. DNA transformation: The process of getting functionally active DNA into cells. Experiment : Purified DNA from Type S (smooth colony) Strep cells is introduced into Type R (rough colonies) Strep cells - Result: Transformation of Type R cells by Type S DNA alters the "genotype" of R cells, since new genes are introduced into these cells thus altering their genetic constitution. The expression of Type S DNA changes the "phenotype“ of the transformed cells, making their colonies look "smooth S" instead of "rough R.“ Genotype is an organism’s genetic constitution. Phenotype is the observed characteristics of an organism as determined by the genetic makeup and the environment. 6

7. II. Transgenic experiments – know this process DNA is the carrier of genetic information. Transgenic mouse:Transfer of a specific gene into the nucleus of a fertilized egg. knockout mutation : The mutation of a gene that eliminates the gene's function. knockout mouse : mouse carrying knockout mutation QUESTION? How to determine gene function? How to produce mouse models of human genetic disease? ANSWER: Produce a knockout mutation of specific genes in the mouse. 7

8. III. Mutation alters phenotype DNA is the carrier of genetic information. Phenotypic differences between individuals are due to differences between genes. One-third of our genes are polymorphic- differences in the nucleotide sequences Differences occurred by mutation of DNA Genetic differences between individuals can have many clinical implications.( aging, drug metabolism). 8

9. Structure of DNA i ) Structure of the bases, nucleosides, and nucleotides ii) Structure of the DNA double helix iii) Complementarity of the DNA strands 9

10. Purines Adenine = 6-amino purine Guanine = 2-amino-6- oxy purine Hypoxanthine = 6- oxy purine Xanthine = 2,6-dioxy purine 10

11. Pyrimidines Uracil = 2,4-dioxy pyrimidine Thymine = 2,4-dioxy-5- methyl pyrimidine Cytosine = 2-oxy-4- amino pyrimidine Orotic acid = 2,4-dioxy- 6-carboxy pyrimidine 11

12. 12 Thymine (T) Guanine (G)Cytosine (C) Adenine (A) Structures of the bases PurinesPyrimidines 5-Methylcytosine (5mC)

13. 5-Methylcytosine (5mC). A common base modification in DNA results from the methylation of cytosine, giving rise to 5- methylcytosine (5mC). 5mC is highly mutagenic. It is believed that this methylation functions to regulate gene expression because 5-methylcytosine (5mC) residues are often clustered near the promoters of genes in so-called "CpG islands.“ The problem that arises from these methylations is that subsequent deamination of a 5mC results in the production of thymine, which is not foreign to DNA. As such, 5'-mCG-3' sites (or mCpG sites) are "hot- spots" for mutation, and when mutated are a common cause of cancer. 13

14. 14 [structure of deoxyadenosine] Nucleoside Nucleotide

15. 15 Nomenclature Purines adenineadenosine guanineguanosine hypoxanthineinosine Pyrimidines thyminethymidine cytosinecytidine +ribose uraciluridine Nucleoside Nucleotide Base +deoxyribose +phosphate