1. Chapter 8
Microbial Genetics
part B

2. Genetics Study of what genes are. How DNA replicate. DNA DNA
How information is expressed.
Regulation of gene expression
How they carry information.
Changes in the genetic material
Genes and evolution
DNA polymerase
RNA polymerase Translation
Gene (DNA) mRNA Protein
5’ ….ATGCCCTGAAAAGAGTCT…..3’

3. Regulation of Bacterial Gene Expression
The cell conserves energy by making only those proteins needed at a particular time.
Constitutive enzymes are produced at a fixed rate
Enzymes of glycolysis are example
Genes encoded these enzymes – constitutive genes
60 – 80% of cell genes
Other enzymes are produced only as needed
Inducible enzymes
Induction – The process that turns on the transcription of a gene or genes
Repressive enzymes
Repression – the regulatory mechanism that inhibits (turns off) gene expression and decrease the synthesis of enzymes

4. Regulation of Bacterial Gene Expression
Regulatory genes (I) code for regulatory proteins
Often repressor proteins
Repressor proteins then attach to a DNA segment known as the operator (O).
By binding to the operator, the repressor protein prevents the RNA polymerase from creating messenger RNA.
Blocking of gene expression is called repression
Repressor protein
Figure 8.13

5. Regulation of Bacterial Gene Expression
Repressor protein activity depends on the presence or absence of an effector substance.
Induction – When the cells are exposed to the compound (substrate) which has to be processed
Repression - When the cells are exposed to a particular end-product.
Enzymes
Substrate Product
Figure 8.13

6. Regulation of Gene Expression - Induction
I – Regulatory gene – encode repressor protein - active; effector substance – substrate
Lac operon is called inducible operon
The lac operon is an operon required for the transport and metabolism of lactose in Escherichia coli and some other enteric bacteria
Effector substance – Lactose
When Lactose is absent:
- Repressor is active
- No transcription
When Lactose is present into the cell:
- It is converted to allolactose (inducer)
Repressor is inactive
- Active transcription – mRNA is synthesize
Induced by presence of Lactose (substrate)
Figure

7. Regulation of Gene Expression - Repression
I – Regulatory gene – encode repressor protein – inactive; Effector substance – product
Tryptophan operon is repressible operon
Trp operon encodes the genes for the synthesis of tryptophan
Effector substance – Tryptophan
Absence of tryptophan
- Repressor is inactive
- Transcription is on
Presence of tryptophan
Tryptophan (corepressor) binds with the repressor protein – active repressor
No transcription
Repressed by presence of Tryptophan ( product)
Figure

8. Regulation of Gene Expression - Catabolite repression
Lactose (disaccharide)  hydrolysis  glucose + galactose
Regulation of Lac operon also depend on the level of glucose in the medium
Glucose concentration in the medium controls the intracellular level of the small molecule cyclic AMP (cAMP), when glucose is no longer available, cAMP accumulates in the cell.
Glucose effect or catabolite repression – inhibition of the metabolism of alternative carbon sources by glucose
Figure 8.15

9. Regulation of Gene Expression: Catabolite repression
CAP – catabolite activator molecule
Lactose present
Glucose is absent
cAMP level is high
Transcription is stimulated
Lactose present
Glucose present
cAMP level is low
Transcription is not stimulated

11. Regulation of Gene Expression - Epigenetic Control
Epigenetic control of gene expression - the switching on and switching off of genes.
Methylating certain nucleotides – turned gene expression off
Methylated (off) genes are passed to offspring cells
Not permanent
Signals from the outside can work through the epigenome to change a cell's gene expression
Contributes to cellular differentiation and development
Aberrant epigenetic control contributes to disease (particularly to cancer)
Biofilm behavior 11

12. Regulation of Gene Expression – Post-transcriptional control
MicroRNAs control a wide range of activities in cells.
Transcription of
miRNA occurs.
miRNA binds to target
mRNA that has at least six
complementary bases.
mRNA is degraded.
DNA
miRNA
mRNA

13. Changes in the genetic material
1. Mutation
A randomly derived change to the nucleotide sequence of the genetic material of an organism.
2. Genetic Transfer and Recombination
Genetic recombination refers to the exchange between two DNA molecules.
It results in new combinations of genes on the chromosome.

14. Mutation Mutation occur spontaneously – spontaneous mutation
Mistakes of DNA polymerase
Genetic recombination
Mutation rate is the probability that a gene will mutate when a cell divides; the rate is expressed as 10 to a negative power.
Spontaneous mutation rate = 1 in 109 replicated base pairs (frequency – 10-9 ) or 1 in 106 replicated genes (10-6 )
Mutations usually occur randomly along a chromosome.
A low rate of spontaneous mutations is beneficial in providing the genetic diversity needed for evolution.

15. Mutation Based on the changes in DNA sequence mutations may be:
Base substitution
Deletion
Insertion
Based on the phenotypic effect on the organism mutations may be:
Neutral
Beneficial
Harmful
Organisms have mechanisms such as DNA repair to remove mutations
Mutation is generally accepted by biologists as the mechanism by which natural selection acts
Generating advantageous new traits – offspring survive and multiply
Generating disadvantageous traits - offspring tend to die out

16. Base substitution (point mutation)
Missense mutation - Change in one base result in change in amino acid
Nonsense mutation - Change in one base results in a nonsense codon
Figure 8.17a, b

17. Mutation – deletion and insertion
One or few nucleotide pairs are deleted or inserted in the DNA
Frameshift mutation – shift the “translational reading frame”
Figure 8.17a, d

18. Identifying mutants - Replica Plating
Positive (direct) selection
Negative (indirect) selection
Example: an auxotrophic mutant cannot synthesize Histidine
100 colony
Figure 8.21

19. Mutagens
Mutagens are agents in the environment that cause permanent changes in DNA.
Mutagens increase mutations to 10–5 or 10–3 per replicated gene
1. Chemical mutagens – Nitrous acid (alters Adenine), nucleoside analogs, benzopyrene
2. Ionizing radiation (X rays and gamma rays) causes the formation of ions that can react with nucleotides and the deoxyribose-phosphate backbone.
3. UV radiation causes thymine dimers
Light-repair separates thymine dimers
Photolyases – enzymes that can
repair UV-induced damage

20. The Ames Test for Chemical Carcinogens
Many mutagens have been found to be carcinogens
Ames test –
Mutated Salmonella his- (lost ability to synthesize histidine)
Mutagenic substance may cause new mutation that reverse the original mutation to his+ ( back mutation or reversions)
Incubation with mutagen / Control – without mutagen
Liver extract – supply all necessary activation enzymes
Figure 8.22

21. Genetic Transfer and Recombination
Genetic recombination,
The rearrangement of genes from separate groups of genes
A molecule of nucleic acid is broken and then joined to a different one
It contributes to genetic diversity.
Vertical gene transfer occurs during sexual reproduction
between generations of cells
Horizontal gene transfer in bacteria
involves a portion of the cell’s DNA being transferred from donor to recipient between cells of the same generation

22. Genetic Recombination
In eukaryotes, recombination occurs in meiosis - chromosomal crossover - exchange of genes between two DNA molecules
Crossover occurs when two chromosomes break and rejoin
Figure 8.23 22

23. Genetic Recombination
When some of the donor’s DNA has been integrated into the recipient’s DNA, the resultant cell is called a recombinant
Donor
Recipient
Recombinant cell
Figure 8.25 23

24. Genetic transfer 1. Transformation
During this process, genes are transferred from one bacterium to another as “naked” DNA in solution.
This process was first demonstrated in Streptococcus pneumoniae and occurs naturally among a few genera of bacteria.
Figure 8.24

25. Genetic transfer 2. Conjugation Requires contact between living cells.
Genetic donor cell is an F+;
F+ cells contain plasmids called F factors;
F factors are transferred to the F– cells during conjugation.
Recipient cells are F–
Converted to F+
Figure 8.27a

26. Conjugation
When the plasmid becomes incorporated into the chromosome, the cell is called an Hfr (high frequency of recombination) cell.
During conjugation, an Hfr cell can transfer chromosomal DNA to an F– cell.
Usually, the Hfr chromosome breaks before it is fully transferred
Figure 8.27b

27. Plasmids
Plasmids are self-replicating circular molecules of DNA carrying genes that are not usually essential for the cell’s survival.
Conjugative plasmids
Carries genes for sex pili and transfer of the plasmid
Dissimilation plasmids
Encode enzymes for catabolism of unusual compounds
Pathogenicity Plasmids
Carrying genes for toxins or bacteriocins
R factors
Encode antibiotic resistance
Plasmids usually occur naturally in bacteria, but are sometimes found in eukaryotic organisms (e.g., the 2-micrometre-ring in Saccharomyces cerevisiae)

28. Genetic transfer
3. Transduction by a Bacteriophage (viruses that infect bacteria).
In this process, DNA is passed from one bacterium to another in a bacteriophage and is then incorporated into the recipient’s DNA. . 2
In generalized transduction, any bacterial genes can be transferred.
Specialized transduction 3 4 5 6
Figure 8.28

29. Transposons
Segments of DNA that can move from one region of DNA to another
Contain insertion sequences for cutting and resealing DNA (transposase)
Complex transposons carry other genes
Figure 8.30a, b

30. Phenotypic effect of mutation
Neutral
Beneficial
Appear new quality
Change metabolic requirement
Loose the ability to metabolize some substrate
Increase activity of some enzyme
Harmful
Lethal effect

31. Evolution
Evolution is the process of change in the inherited traits of a population of organisms from one generation to the next
. 1. Diversity is the precondition for evolution
Genetic mutation and recombination provide a diversity of organisms
2. The process of natural selection (positive selection)
Allows the growth of those best adapted to a given environment

32. Learning objectives
Explain the regulation of gene expression in bacteria by induction, repression, and catabolite repression.
Classify mutations by type, and describe how mutations are prevented and repaired.
Define mutagen
Describe the effect of mutagens on the mutation rate.
Compare the mechanisms of genetic recombination in bacteria.
Differentiate between horizontal and vertical gene transfer.
Describe the functions of plasmids and transposons.
Outline methods of direct and indirect selection of mutants.
Discuss how genetic mutation and recombination provide material for natural selection to act on.