Division of bacterial cells takes about 20-25 minutes.

1. Genetic Variation = refers collectively to the diversity of genomes that give rise to different individuals • includes small differences between genomes of members of the same species and greater differences between the genomes of different species • the ultimate source is changes in DNA (mutations) that alter its nucleotide sequences • some small due to rare “mistakes” in DNA replication and repair • some large, due to different processes including DNA recombination, the activities of viruses, and mobile genetic elements. • Also sexual reproduction results in re-assortment of the gene pool into new combinations in different individuals

2. Division of bacterial cells takes about 20-25 minutes. With no mistakes, each parent gives rise to identical daughters Can be grown in liquid medium with nutrients and oxygen at around 37˚C

3. Grown on solid surface of agar, colonies develop from individual bacteria if the original suspension is dilute, and spread over the surface of the agar. Individual colonies can be selected for study.

4. Mutations in bacteria can be selected by a change in the environment. Grown in the presence of rifampicin, mutants with an RNA polymerase that is not disabled by this antibiotic survive and take over the culture. In any large population of bacteria there will be many different “mutants” (genetic variants). These can be selected for during a change in the environment (natural selection).

5. Bacterial genes can be acquired from other bacteria by several different routes. Direct cell to cell transfer during bacterial mating or conjugation is shown here. An example of DNA exchange: E. Coli strain with mutation making it unable to make methionine grown with E. Coli strain with mutation making it unable to make leucine results in the appearance of a new strain able to make both. How? DNA recombination Quesstions 2,3,5,6, 8,9, 11,13,14,17

6. Bacterial mating occurs between a bacterium with an F plasmid and one without one. A cytoplasmic bridge is formed, DNA is duplicated and transferred. 1. 2. Plasmids are small, circular, double-stranded DNA separate from the bacterial chromosome which can replicate itself. F plasmids (fertility) contain the genes needed for conjugation.

7. Usually only a small number of genes on the plasmid are transferred. Rolling circle replication

8. Transfer of bacterial chromosomal genes by an F plasmid after integration into the bacterial chromosome. Widespread use of antibiotics leads to many clinical isolates that are resistant to the antibiotic typically used – as in Neisseria gonorrhoeae to penicillin. These contain plasmids that encode proteins that inactivate the antibiotics

9. Bacteria can take up the DNA from their surroundings - Bacterial Transformation. Uptake of naked DNA is active and receptor-mediated and has become an important tool in the laboratory for research and biotechnology.

10. DNA aligns so that base pairs of homologous sequences match complementarily Site of exchange can occur anywhere in this region Cleavage of the DNA molecules and rejoin is precise, no nucleotides are lost or gained Once inside of a bacteria, naked DNA will only survive and be passed on to progeny if it is incorporated into the bacterial chromosome. Homologous Recombination occurs in all organisms and takes place between two DNA molecules with similar sequences.

13. Cross-strand exchange or Holliday junction

14. Homologous recombination can lead either to an exchange of DNA between two molecules or to the combination of two circular DNA molecules into one. F plasmid contains a short homologous stretch of DNA Cells utilize specialized proteins to facilitate homologous recombination; these proteins nick DNA, catalyze strand exchange, and cleave Holliday structures.

15. Genes can be transferred between bacteria by bacterial viruses - bacteriophages. This is a simple virus - simpler than any real virus. Consists of a small double-stranded DNA moleculs must enter the cell, replicate its DNA, and transcribe some, especially the protein coat. Viral DNA and protein coat spontaneously assemble.

16. Encodes 50 - 60 proteins Bacteriophage lambda Lytic infection Latent state Prophage lies dorman Damage to bacteria, ultraviolet light

17. Integrates by Site-specific recombination Virus encodes a specialized enzyme - lambda integrase - which recognizes a specific sequence of bacterial DNA and a specific sequence of viral DNA. It binds these and then brings them together, breaks both, and then rejoins them Exits by doing the same thing in reverse. Excises itself inaccurately on occasion The modified lambda infects a new host. Two ways the bacterial DNA is transferred to another bacteria. 1. Integration 2. Homologous recombination (transduction)

18. Homologous recombination can lead either to an exchange of DNA between two molecules or to the combination of two circular DNA molecules into one.

19. Transposable Elements(transposons) create more genetic diversity. Range from several hundred to tens of thousands of base pairs. Typical lab E. coli contains 10-20 different transposons, with many having multiple copies. Transposons move within a DNA molecule by using a special recombination enzymes - transposases - encoded by the transposon. Sequences in red are recognized only by the particular transposase encoded. Transposons can “pick up” host DNA by homologous recombination between two identical transposons.This probably led to the antibiotic resistant plasmids and transposons.

20. Transposons provide a number of advantages: 1. Some contain transcriptional promoters, land near a bacterial gene, and cause it to be expressed at a different level or under different controls. 2. Some are present in multiple copies, allowing rearrangement by homologous recombination between transposons. 3. Some contain drug resistant genes which then can jump into plasmids and be transmitted to other bacteria Donor DNA and targe can be on the same molecule or on different molecules (bacterial genome + plasmid) Generally a transposon can use only one of these methods.

21. DNA replication is remarkably accurate. • However, genomes are in a state of slow but constant change • mutations accumulate over time • genes are transferred from one bacterium to another • viral genomes move in and out • transposons change position • Most alterations in the genome are harmful, but, when the environment changes (antibiotics are present) genetic variation is crucial for survival.

22. Sources of Genetic change in Eucaryotic Genomes • Bacterial genomes are streamlined, genes are closely packed with relatively little spacer DNA and few introns • Mammalian genome contains enormous amounts of non-gene DNA including introns, spacer DNA between genes and various types of repetitive DNA sequences with similarities to transposons. Also a large amount of gene duplication, leading to the large families of highly related genes and different forms of each gene - alleles. • this DNA has far-reaching effects • alleles increase the “gene pool” - the entire collection of alleles of every gene present in a species

23. Random DNA duplications create families of related genes. Ex. Different forms of actin expressed in different types of muscle cells and in other cell types. There are 5 beta-globin genes, each produced at a different time during embryonic, fetal, and adult development. Each with a different oxygen-binding and -releasing characteristic appropriate for the time it is expressed. And each under independent regulation. Comparing the sequences of genes like the beta-globin gene leads to models like this which may reflect the evolution of thegene through thousands of years.

24. Gene duplication by unequal crossing-over.

25. Genes encoding new proteins can be created by recombination of exons, without damaging the exon, due to the presence of introns. New mRNA can be spliced correctly due to the presence of the proper intron sequences.

26. The presence of introns greatly increases • the probability that DNA duplications will give rise to functional genes • and the probability that a chance recombination event can generate a functional hybrid gene by joining two initially separate exons coding for different protein domains – exon shuffling.

28. Some Results of exon shuffling through time.

29. A large part of the DNA of multi-cellular eucaryotes consists of repeated, non-coding sequences • About 70% in humans is “unique” DNA coding for proteins or RNA and including intron DNA • Remaining 30% includes two types of repeated, non-coding sequences • Satellite DNA = highly repeated short sequences that form serial arrays (about 1/3), which can be clustered at the centromeres and at the ends of chromosomes. • Function is unknown and it varies greatly between individuals of the same species • Repetitive sequence DNA composed of complex repeated sequences interspersed throughout the genome derived from a few types of transposable DNA sequences like those in bacteria • DNA in humans and primates is unusual in that it contains a remarkable number of two of these transpoable DNA sequences.

30. About 10% of the human genome consists of two families of transposable sequences. Some move using the cut-and-past mechanism in Figure 18A. Others move via an RNA intermediate = retrotransponons – perhaps unique to eucaryotes. L1 transposable element, or LINE-1 = 4% of the total mass of the human DNA The reverse transcriptase is encoded in L1. Alu sequence is shorter and is about 5%. It does not code for a reverse transcriptase – requires one encoded elsewhere. Many are non-movable

31. Evolution of genomes has been accelerated by transposable elements. They are a significant source of mutation Insertion into coding sequences of a gene or into its regulatory region is relatively frequent cause of spontaneous mutations in some organisms. Ex. Some mutations in Factor VIII (hemophilia) result from insertion of transposable elements into the gene. They provide opportunities for genome rearrangements by serving as target of homologous recombination.

32. Exon shuffling caused by a transposon. Transposase uses the beginning of the 1st element and the end of the 2nd element to replicate and move the segment, including exon 2A into a different gene.

33. DNA rearrangement caused by a transposable element can produce a dramatic change in the organism. normal Legs where antennae should be

34. Lytic effect of virus

35. Genomes of viruses can vary greatly. The amount of DNA or RNA that can be packaged is limited. Viral genomes encode few proteins and rely on host enzymes to replicate and translate their genes, including viral coat proteins and proteins that attract host enzymes to replicate their genome.

36. Viruses are parasites that can reproduce themselves only inside a living cell. This is the life cycle of a single-stranded RNA virus. RNA viruses must encode an RNA replicase, to replicate its genome. Negative stranded RNA viruses must carry an RNA replicase in their protein coat.

37. Viruses are parasites that can reproduce themselves only inside a living cell. The smallest viruses contain 3 genes. protein coat is made up primarily of one polypeptide. More complex viruses have genomes up of up to several hundred genes and are surrounded by an elaborate shell composed of many different proteins.

38. HIV is a retrovirus Latent = problem for treatment Host V Integrase V

39. Since reverse transcriptase is not used by host cells it is a prime target of drug development Retroviruses may have derived from retrotransponsons that long ago acquired additional genes encoding coat proteins etc. Viruses that integrate into host cell DNA are, like transposable elements, potential agents of genetic change.

40. About 10% of the human genome consists of two families of transposable sequences. Some move using the cut-and-past mechanism in Figure 18A. Others move via an RNA intermediate = retrotransponons – perhaps unique to eucaryotes.

41. Retroviruses can pick up host genes. This can lead to cancer. The virus is then known as a tumor virus. Ex. Rous sarcoma virus picked up a gene from chickens that encodes a protein kinase important in control of cell division. This gene was altered in the virus to become hyperactive. Expression of the viral src gene (an oncogene) leads to the loss of control of cell division - cancer.

43. Egg with sperm bound Sexual reproduction is costly in terms of resources spent but is advantageous in terms of genetic variation created and changing environments.

44. Genetic variation

45. Genetic variation chiasmata

47. Reassortment = genetic variation 23 possibilities 2n possibilities