1. Chapter 10
Prokaryotic Genetics

2. Plasmids
Plasmids: genetic elements that replicate independently of the host chromosome
Small circular or linear DNA molecules
Range in size from 1 kbp to > 1 Mbp; typically less than 5% of the size of the chromosome
Carry a variety of nonessential, but often very helpful, genes
Abundance (copy number) is variable

4. Prokaryotic diversity
Why are prokaryotes so diverse when they do not reproduce sexually?
Mutation
Inherited change in genotype
Small gradual change
Recombination
Integration of DNA (from another organism or genetic element) into chromosomal DNA
Sometimes very large changes

5. Mutants Some of this we already covered in chapter 6 Often silent
Change in nucleic acid, but no change in amino acid coded for
Sometimes phenotypic changes
Requires change in amino acid
Lethal, neutral, beneficial
Sometimes a change in amino acid (so not silent) but no change in protein so no phenotypic change

6. Point mutations Involving one base pair Nucleic acid base substitution
Missense = changes the codon (1st or 2nd base)wrong amino acid
Changes protein
Sometimes a phenotypic change and sometimes not
Nonsense= changes the codon and codes for a stop codon
Translation terminated early protein often non-functional
Silent= changes last base in codonsame amino acid usually
Degeneracy of the code

7. Point mutation: base substitution

8. Point mutations Transitions Transversions
One purine base (A or G) is substituted for another purine or one pyrimidine base (C or T) is substituted for another pyrimidine
Transversions
A purine is substituted for a pyrimidine or a pyrimidine is substituted for a purine

9. Point mutations Frameshift mutation
Insertion or deletion of a few nucleotides causing a reading frame shift and disruption of translation
Insertion= +1 frameshift and deletion = -1 frameshift

10. Point Mutation: Frameshift

11. Other Mutations: More Bases
Large deletions: more likely lethal
Can only be restored by recombination
Large insertions: often inactivate gene
Can only be reverted by large deletion
Translocations: movement of a large segment from one area to another (ex. Transposons)
Inversion: orientation of DNA reversed

12. Wild type versus mutant
hisC gene codes for HisC protein
Mutation in the hisC gene are called hisC1, hisC2 etc.
=genotype
Phenotype: His + or His –
His+ - capable of making histidine
His – not capable of making histidine

13. Isolation of mutants
Selectable mutants: can select for a phenotype by subjecting population to a selection factor
Selectable– antibiotic resistance
Only certain bacteria will grow on a particular antibiotic
We will do a transformation lab where we will grow bactiera on certain antibiotics
Used for cloning
Non-selectable – loss of color (may still have a selective advantage in a natural ecosystem, but cannot easily select for the trait in culture)
Non-selectable mutants have to be screened
Some will have a different color but all will grow

14. Isolation of mutants His C – mutant: cannot make histidine (auxotroph)
Auxotroph: a nutritional mutant (requires a growth factor that the WT parent did not require)
Prototroph: the WT parent from which the auxotroph was derived
Replica plating is one method to screen for nutritional mutants

15. Replicate plating to isolate auxotrophic mutants: grow with His but not without His

16. Molecular Basis of Mutation
Induced mutations
Those made deliberately
Spontaneous mutations
Those that occur without human intervention
Can result from exposure to natural radiation or oxygen radicals
Point mutations
Mutations that change only one base pair
Can lead to single amino acid change in a protein or no change at all

17. Types of mutagens Chemical
Nucleotide base analogs: faulty base pairing

18. Types of mutagens Radiation
Nonionizing: causes pyrimidine dimers, which causes problems with replication and transcription
EX) UV light
Ionizing:
More energy
Penetrates through glass
Free radicals will damage DNA and disrupt base pairing
X-rays and Gamma radiation

20. Genetic recombination
Physical exchange of DNA between genetic elements
Homologous recombination
Process that results in genetic exchange between homologous DNA from two different sources

21. A Simplified Version of Homologous Recombination

22. Genetic exchange in prokaryotes
Donor DNA is transferred to recipient cell in 3 possible ways
Transformation: free DNA released from one cell is taken up by another
Transduction: DNA transfer is mediated by a virus
Conjugation: plasmid transfer with cell to cell contact

23. Mechanisms of Transformation in Gram-Positive Bacteria

24. Transduction
Transfer of DNA from one cell to another is mediated by a bacteriophage
Generalized transduction: DNA derived from virtually any portion of the host genome is packaged inside the mature virion

25. Generalized transduction

26. Conjugation: Genetic transfer involving cell to cell contact
Donor cell
contains a conjugative plasmid
Produces a sex pilus
F plasmid produces F pilus
Pili make contact with recipient cell and pull it closer
Only donor cells produce pili

27. Transfer of Plasmid DNA by Conjugation

28. Mobile DNA: Transposable Elements
Discrete segments of DNA that move as a unit from one location to another within other DNA molecules (i.e., transposable elements)
Transposable elements can be found in all three domains of life
First observed by Barbara McClintock