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Chapter 8: Bacterial Genetics. Important Point:. If you are having trouble understanding lecture material: Try reading your text before attending lectures. And take the time to read it well!. Bacterial Genetics.
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Important Point: If you are having trouble understanding lecture material: Try reading your text before attending lectures. And take the time to read it well!
Bacterial Genetics • “Acquiring genes through gene transfer provides new genetic information to microorganisms, which may allow them to survive changing environments.” • “The major source of variation within a bacterial species is mutation.” • “In mutations, usually only a single gene changes at any one time.” • “In contrast, gene transfer results in many genes being transferred simultaneously, giving the recipient cell much more additional genetic information.”
Most bacteria are haploid which means that there is no such thing as dominance-recessive relationships among bacterial alleles. • Bacteria don’t have sex in the animal/plant sense of sex (i.e., mating followed by recombination of whole genomes). • Instead, bacteria acquire DNA from other bacteria through three distinct mechanisms: • Transformation • Transduction • Conjugation • This DNA may or may not then recombine into the recipient’s genome. • We use phrases like “Lateral” or “Horizontal” Gene Transfer to describe these sexual interactions. • Bacterial DNA is also subject to mutation, damage (not the same thing as mutation), and natural selection. Bacterial Genetics Overview
Wild Type refers to the microorganism as isolated from the wild. • A mutated microorganism that has lost a metabolic function, particularly an ability to synthesize a specific growth factor, is called an Auxotroph. • The wild-type parent to an auxotroph is called a Prototroph. • A Mutation is found in a gene; a mutant is an organism harboring a Mutation. • We designate mutant phenotypes using three-letter abbreviations; the phenotype of a tryptophan-requiring auxotroph would be described as Trp-. • A bacterium that has mutated to resistance to an antibiotic (or other substance) is given the superscript “R”; thus, the phenotype ampicillin resistance is indicated as AmpR. • Mutants can be spontaneous or induced by a Mutagen; an agent that causes DNA to mutate. Mutation: Terms & Concepts
Base Substitution • Point mutation = single base is substituted. • Missense mutation = base change changes single amino acid to different amino acid. • Nonsense mutation = base change changes single amino acid to stop codon. • Null or Knockout mutation = mutation that totally inactivates a gene. • Deletion or insertion mutation = change in number of bases making up a gene. • Frameshift mutation = insertion or deletion of something other than multiples of three bases. • Frameshifts typically radically change downstream codons, generating stop codons, and typically knocking out gene function. • Reversion mutation = mutated change back to that of wild type. Types of Mutations
The mutation rate of different genes usually varies between 10-4 and 10-12 mutations per cell division (essentially equivalent to per cell). • 10-4 = one in 10,000; 10-12 = one in one trillion. • To calculate the probability of two independent mutations we multiple the two mutation rates. • Thus, if streptomycin resistance occurs at a rate of 10-6 mutations per cell division and the rate of mutation to resistance to penicillin is 10-8 then the rate of mutation to both antibiotics is 10-6 * 10-8 = 10-14 (note that the exponents are added). • That is, we would have to have a population of one-hundred trillion cells to have one double mutant, which even for bacteria is a lot of cells. • This is the basis for Combination Therapy, e.g., the use of more than one chemotherapeutic against tuberculosis, HIV, cancer, etc. • The odds of sufficiently multiply resistant mutants drops with each new chemotherapeutic added to the mix. Rates of Mutation
Indirect Selection:Isolation of ts Mutants This is one example of isolation of mutants carrying conditionally lethal mutations found in essential genes.
DNA-Mediated Transformation Note that DNA is taken up naked from the environment.
Original Transformation Exp.F. Griffith (1928) using pneumococci
Artificial Competenceby Electroporation Competence denotes the ability to take up DNA naked from the environment. Most bacteria are not naturally competent but many can be made artificially so. Artificially induced competence is very important to gene cloning.
Generalized Transduction Bacteriophages are viruses that only infect (and can kill) bacteria.
F plasmids encode genes that allow both their replication and transfer. • They are thus known as Self-Transmissible Plasmids. • There are other plasmids that can take advantage of conjugation but don’t encode the the necessary genes. These are non-self transmissible plasmids. • Transduction is also capable of transferring smaller plasmids. • R plasmids are named not for their mode of transmission but instead for the resistance genes that they encode such as to antibiotics. • Some plasmids are present in bacteria in low copy numbers (1 or 2/bacterium) whereas other plasmids are present in high copy numbers (such 100s/bact.). • Plasmids, R and otherwise, can have very wide host ranges allowing easy transfer of already evolved genes between bacterial species. F and Other Plasmids
Self-Transmissible R Plasmid Note multiple resistance genes. Resistance Transfer Factor (conjugation genes)
Transfer of non-R Virulence Factors • Genes that can make bacteria more virulent (able to cause disease) are called Virulence Factor genes. • Virulence factors include fimbriae that allow attachment to host tissues, exotoxins, etc. • Virulence factor genes may be transferred by transformation, transduction, or conjugation. • Virulence factor genes tend to congregate on bacterial chromosomes in regions known as Pathogenicity Islands. • New bacterial pathogens can emerge via the uptake of entire pathogenicity islands transferred intact from unrelated bacteria.
Not all incoming DNA is necessarily good for the receiving bacterium (i.e., DNA can be parasitic). • Bacteria employ Restriction Enzymes to protect themselves from the foreign DNA. • Restriction enzymes recognize specific, palindromic (same spelling backward and forward) DNA sequences of 4 to 8 base pairs in length that are known as Recognition Sequences. • Bacteria also employ Modification Enzymes that modify DNA to protect it from Restriction Enzymes. • Together these are called Restriction-Modification Systems. • Restriction enzymes are crucial components of genetic engineering. Transfer Protection: R-M Systems
Restriction Endonuclease Action Note in particular that DNA is cut at palindromic regions.
