1. © 2004 Wadsworth – Thomson Learning Chapter 6 The Genetics of Microorganisms

2. © 2004 Wadsworth – Thomson Learning Structure of DNA Two strands Nucleotides –Hydrogen bonds between strands –Neighboring deoxyribose connected 3’ of one deoxyribose to 5’ of next deoxyribose Phosphate in between Double helix Base pairing –G and C –A and T Figure 6.1

3. © 2004 Wadsworth – Thomson Learning Roles of DNA Replication –cell division –need accurate copy Gene expression –DNA –RNA –Protein Figure 6.2

4. © 2004 Wadsworth – Thomson Learning DNA Replication Semi-conservative –old strand-template –new strand- complementary Replication fork –multiple enzymes –DNA unwinds –exposes nucleotides –synthesize new strand –one direction: 5’ to 3’ Figure 6.3

5. © 2004 Wadsworth – Thomson Learning DNA Replication Complementary nucleotides match (A=T; G=C) DNA polymerase III binds nucleotides releasing pyrophosphate Figure 6.3

6. © 2004 Wadsworth – Thomson Learning Bacterial chromosomes Replication of circular chromosome Origin of replication –bubble forms –DNA unwinds Replication occurs in both directions Two replication forks Continues until replication forks meet Strands separate Figure 6.4

7. © 2004 Wadsworth – Thomson Learning Replication fork Figure 6.5

8. © 2004 Wadsworth – Thomson Learning DNA Replication Leading strand –replication is continuous (5’ to 3’) primase makes primer DNA added to primer fork opens and replication continues Lagging strand –polymerization in only one direction can’t go 3’ to 5’ –short segments synthesized (Okazaki fragments) when fork opens, new primer is made synthesis in direction away from fork fragments are joined together by DNA ligase

9. © 2004 Wadsworth – Thomson Learning Transcription RNA polymerase binds DNA at site of promoter Figure 6.6

10. © 2004 Wadsworth – Thomson Learning Transcription DNA unwinds nucleotide bases are exposed Figure 6.6

11. © 2004 Wadsworth – Thomson Learning Transcription ribonucleotides pair with exposed bases –uracil in RNA replaces thymine –U binds A ribonucleotides are polymerized into growing RNA chain Figure 6.6

12. © 2004 Wadsworth – Thomson Learning Transcription Termination sequence Release of transcript single strand RNA DNA closes Figure 6.6

13. © 2004 Wadsworth – Thomson Learning Transcription Role of RNA from transcription –mRNA template which encodes the protein –tRNA transfer amino acids used to build the protein –rRNA part of ribosome which is the site of protein synthesis All used for translation

14. © 2004 Wadsworth – Thomson Learning Translation Production of proteins Based on genetic information of DNA Genetic code –Codon has three nucleotide –Four different nucleotides –64 possible combinations –20 amino acids Redundancy Nonsense codons Figure 6.8

15. © 2004 Wadsworth – Thomson Learning Translation tRNA –binds an amino acid specific amino acid for each tRNA –Anticodon recognizes codon three nucleotide sequence in mRNA which encodes a specific amino acid –activated with ATP Figure 6.7

16. © 2004 Wadsworth – Thomson Learning Translation Ribosome binds to mRNA –specific region –start codon Methionine –Ribosome binding region Shine-Dalgarno sequence Figure 6.9

17. © 2004 Wadsworth – Thomson Learning Translation tRNA with appropriate anticodon and specific amino acid binds to the codon on the mRNA –A site second tRNA binds in similar fashion –P site two amino acids are joined in a peptide bond Figure 6.9

18. © 2004 Wadsworth – Thomson Learning Translation Ribosome moves along mRNA first tRNA without amino acid is removed second tRNA with both amino acids moves to P site Figure 6.9

19. © 2004 Wadsworth – Thomson Learning Translation New tRNA enters A site Growing amino acid chain is transferred to new amino acid Figure 6.9

20. © 2004 Wadsworth – Thomson Learning Translation steps repeat –ribosome moves –one codon at a time protein chain –one amino acid added for every codon Figure 6.9

21. © 2004 Wadsworth – Thomson Learning Translation Continues until nonsense (stop) codon is reached no tRNA matches ribosome is removed protein chain is released Figure 6.9

22. © 2004 Wadsworth – Thomson Learning Transcription and Translation Simultaneous transcription and translation mRNA chain is transcribed translation begins multiple ribosomes on single mRNA –polysome Figure 6.10

23. © 2004 Wadsworth – Thomson Learning Regulation of genes Transcription –Production of regulatory proteins Bind DNA near the promoter Example: Lactose operon –Interruption of transcription Attenuation Translation –Ribosomal proteins Global regulation –Catabolite repression –Nitrogen regulation –Phosphorus regulation –Stringent response –Heat shock proteins

24. © 2004 Wadsworth – Thomson Learning Transcriptional regulation lac operon lacZ lacY lacA –regulated by lacI Lactose absent –repressor binds –stops transcription Figure 6.11

25. © 2004 Wadsworth – Thomson Learning Transcriptional regulation Lactose present –repressor bound by product of lactose allolactose –transcription occurs –gene products of all genes are made Figure 6.11

26. © 2004 Wadsworth – Thomson Learning Attenuation Histidine operon –Histidine present –Leader protein made Translation occurring simultaneously with transcription Requires histidine –Attenuator loop forms on mRNA Displaces RNA polymerase Stops transcription Figure 6.12a

27. © 2004 Wadsworth – Thomson Learning Attenuation Histidine absent –Leader protein not made Not enough histidines to complete protein –Antiterminator loop forms Prevents attenuator loop from forming RNA polymerase continues Figure 6.12b

28. © 2004 Wadsworth – Thomson Learning Regulation of translation Expression of ribosomal proteins –Unused proteins bind to encoding mRNA –Inhibit translation Figure 6.13

29. © 2004 Wadsworth – Thomson Learning Two component regulation Phosphorylation of sensor Phosphate passed to response regulator Response regulator reacts with DNA changing gene expression –Increase –decrease Figure 6.14

30. © 2004 Wadsworth – Thomson Learning Genetic Information Genome –total DNA of a cell –most have single circular chromosome –some have linear chromosome Plasmids –small, circular, extrachromosomal DNA encode beneficial factors resistance factors (antibiotic) conjugative plasmids –transfer to other cells Genotype: genetic makeup Phenotype: appearance and function

31. © 2004 Wadsworth – Thomson Learning Changes in Genetic Information Mutations –chemical change in DNA chemical mutagens –Bind DNA –Change in DNA physical mutagens –UV light –Ionizing radiation biological mutagens –Transposable elements »Insertion sequences »transposons Figure 6.16

32. © 2004 Wadsworth – Thomson Learning Consequence of mutations Types of mutations –base substitution wrong nucleotide –deletion mutation nucleotides deleted –Inversion reverses order of a segment –Transposition moves a segment of DNA –Duplication identical new segment Results –Lethal mutation –Conditional expressed mutations Figure 6.15

33. © 2004 Wadsworth – Thomson Learning Physical mutagen--UV damage UV light –stimulates neighboring bases to form dimers thymine dimers –activate repair systems Figure 6.17

34. © 2004 Wadsworth – Thomson Learning Physical mutagen--UV damage Thymine dimers distort the DNA structure Enzymes remove the damaged nucleotides Figure 6.17

35. © 2004 Wadsworth – Thomson Learning Physical mutagen--UV damage Repairs may result in incorrect nucleotide replacement Mutation is result Figure 6.17

36. © 2004 Wadsworth – Thomson Learning Selecting and identifying mutants Direct selection –Conditions favor growth of desired mutant –Growth of bacteria in presence of antibiotic –Only successful growth are mutants Indirect selection –Prevent growth of mutant –Kill growing cells –Desired mutants larger percentage of population –Isolate mutants Site-directed mutagenesis –Recombinant DNA manipulation

37. © 2004 Wadsworth – Thomson Learning Selecting and identifying mutants Brute strength –Screen large numbers –Replica plating Transfer large numbers of colonies Track growth Figure 6.18

38. © 2004 Wadsworth – Thomson Learning Ames Test Figure 6.19

39. © 2004 Wadsworth – Thomson Learning Transformation DNA exits one cell, taken up by another cell –Natural few bacteria take up DNA fragments –Artificial--induced in laboratory useful tool for recombinant DNA technology Figure 6.20

40. © 2004 Wadsworth – Thomson Learning Conjugation Conjugative plasmids –plasmids transfer –genetically encoded –F plasmid in E. coli –sex pilus connect two cells one cell F + one cell F - Figure 6.21

41. © 2004 Wadsworth – Thomson Learning Conjugation –One strand of plasmid DNA is broken (nicked) –replication begins –synthesized linear strand enters F - cell –linear strand is copied forming a complete plasmid –both cells are F + Figure 6.21

42. © 2004 Wadsworth – Thomson Learning Transduction Bacteriophage –virus that infects bacteria –reproduce in bacteria –some phages contain bacterial DNA rare event transducing particle –cell lysis and release normal phage transducing particles Figure 6.23

43. © 2004 Wadsworth – Thomson Learning Transduction –Transducing particles infect other bacteria inject bacterial DNA into new cell –genetic exchange one bacteria cell to another –integration into chromosome Figure 6.23

44. © 2004 Wadsworth – Thomson Learning Eukaryotic Microorganisms Genetic exchange –Similar to plants and animals –Haploid gametes fuse Figure 6.23