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. A-T hydrogen bonding

6. G-C hydrogen bonding

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

8. 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

9. **DNA structure overview** complementary strands (antiparallel) 3 Angstrom separation of 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? supercoiled DNA (DNA-binding proteins)

10. 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

11. Restriction and modification

12. DNA Replication: addition of a nucleotide

13. Semiconservative replication

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

15. DNA Replication

16. Bidirectional replication

17. Okazaki fragments

19. Proofreading by DNA polymerase III

20. 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)

21. DNA Sequencing

22. 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

23. Initiation of Transcription Pribnow box=tataat

24. Transcription

25. Completion of transcription

26. Example of termination sequence

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

28. 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?

29. 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

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

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

32. Growing protein polymer

33. Translocation

34. 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

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