1. Central dogma: Information flow in cells

2. Nucleotides Pyrimidine bases: Cytosine (C), Thymine (T), Uracil (U, in RNA) Purine bases: Adenine (A), Guanine (U)

3. Prokaryotic gene coding

4. Eukaryotic processing of rRNA

5. DNA Replication: addition of a nucleotide

6. DNA duplex formation

7. A-T hydrogen bonding

8. G-C hydrogen bonding

9. 3D structure of DNA

10. Inverted repeats in DNA

11. Formation of Stem-loops

12. “Sticky ends”

13. Hairpins

14. Genetic Elements Prokaryotes: Chromosome, plasmid, viral genome, transposable elements Eukaryotes: Chromosomes, plasmid, mitochondrion or chloroplast genome, viral genome, transposable elements

15. Melting of DNA Melting means separation of two strands from the heteroduplex Melting temperature of DNA is dependent on the relative number of AT and GC pairs Melted DNA can hybridize at temperatures below melting temperature –This process can be used to test relatedness between species (interspecies DNA-DNA hybridization) –It is also possible to reanneal DNA with rRNA to test relatedness of one species rRNA with the rRNA genes of another species

16. Reannealing DNA

17. **DNA structure overview** complementary strands (antiparallel) 3 Angstrom separationof hydrogen bonds sugar phosphate backbone held together with hydrogen bonding between bases size is expressed in nucleotide bases pairs. E. coli has 4600 kbp. (E. coli chromosome is > 1mm, about 500X longer than the cell itself. How can the organism pack so much DNA into its cell? each bp takes up to 0.34nm, and each helix turn is 10bp(or 34 Angstroms), therefore how long is l kb of DNA? and how many turns does it have? inverted repeats, stem-loop, hairpins, sticky ends supercoiled DNA (DNA-binding proteins) relaxed, nicked circular DNA

18. Supercoiled and relaxed DNA

19. DNA Organization In prokaryotes: naked circular DNA with negative supercoiling –Negative supercoiling is introduced by DNA gyrase (topoisomerase II) –Topoisomerase I relaxes supercoiling by way of single- strand nicks In eukaryotes: linear DNA packaged around histones in units called nucleosomes –The coiling around histones causes negative supercoiling

20. Restriction and modification

21. DNA Replication: addition of a nucleotide

22. Semiconservative replication

23. Initiation of DNA replication Origin of replication= oriC = ~300bp Templates, primers, polymerase, primase

24. DNA Replication

25. Bidirectional replication

26. Okazaki fragments

28. Proofreading by DNA polymerase III

29. Replication overview 1. origin of replication+ 300 bases, recognized by specific initiation proteins = replication fork 2. bidirectional, therefore leading and lagging strands helicase unwinds the DNA a little (ATP-dependant) single-strand binding protein prevents single strand from reannealing Primase, DNA polymerase III and DNA polymerase I (also 5' to 3' exonuclease activity), ligase Okazaki fragments Topoisomerases, and supercoiling regulation 3. Proofreading (3 to 5' exonuclease activity by DNA pol III)

30. DNA Sequencing

31. Transcription RNA plays an important role tRNA, mRNA, rRNA Name three differences between chemistry of RNA and DNA RNA has both functional and genetic roles

32. Initiation of Transcription Pribnow box=tataat

33. Transcription

34. Completion of transcription

35. Example of termination sequence

36. More transcription Polycistronic mRNA How can mRNA be used in microbial ecology? Antibiotics and RNA polymerases

37. RNA processing Removal of introns Ribozymes (nobel prize-Tom Cech and Sid Altman) RNA-splicing enzymes Origins of life? Which came first RNA or DNA?

38. The genetic code Notice that the wobble base generally makes minor changes in the amino acid AUG is the start code (formyl methionine) for bacteria UAA, UAG, UGA are stop codons Specific tRNA for each other codon

39. Codon and Anticodon; Wobble

40. tRNA associated with codon ~60 specific tRNAs in prokaryotes

41. mRNA, tRNA and ribosomes Shine Dalgarno sequence GTP and Elongation Factors (EF)

42. Growing protein polymer

43. Translocation

44. Role of rRNA in protein synthesis Structural and functional role 16S rRNA involved in initiation –Base pairing occurs between ribosome binding sequence on the mRNA and a complementary seq on the 16S rRNA 23S rRNA involved in elongation –Interacts with EFs

45. Chaperones (heat-shock proteins)

46. Overview of today Summarized basic DNA structure DNA replication DNA sequencing Transcription RNA processing Translation Role of rRNA in protein synthesis