Chapter 7.

Microbial Genetics
Genes, Genomes and Chromosomes
• Genetics is the study of heredity. DNA is the genetic material of all cellular
organisms (some viruses use RNA). Mendel discovered gene segregation during
reproduction, refuting the previous popular belief that offspring was a "blending"
of parental genetic materials. Adaptation of organisms to their habitats is caused
by Darwinian natural selection, not due to the Larmarckian heritability of
acquired traits.

• The total genetic information in a cell is called genome. A typical bacterial

genome contains a single circular chromosome and plasmids (shorter pieces of
DNA). A eukaryotic genome contains multiple chromosomes as well as DNA in
mitochondria and chloroplasts (in plants). Diploid eukaryotes like human have
both paternal and maternal chromosomes in a cell's genome. In contrast, bacteria
are haploid organisms with only a single copy of chromosome. The size of
bacterial genomes is about one to a few million base pairs (Mb, e.g., 4.5 Mb in
E.coli).

• Genes exist on loci (locations) of a chromosome. In a natural population, a

genetic locus can have multiple versions of the same gene ("alleles"). Alleles,
which are different DNA sequences of the same gene in a natural population, are
results of DNA mutations. Evolution is a process of allele replacements in natural
populations.

Flow of Genetic Information

• DNA replication: one parental DNA gives rise to two identical copies of daughter
DNAs. DNA polymerase is the key enzyme for DNA replication: it has a high
proof-reading capability, to make sure daughter DNAs are an highly accurate
copy of parental DNA (it's error rate is about 1 mis-incorporation of base for
every billion bases). Bacterial chromosome usually have a single origin of
replication.

• Transcription: the synthesis of a complementary strand of RNA (called

messenger RNA, or mRNA) from a DNA template. An mRNA carries the coded
information for making a protein. RNA polymerase is the key enzyme: (a) it
uses a portion of DNA as a template; (b) it recognizes and binds to a specific site
of DNA called "promoter" to start transcription. In bacteria, a group of
functionally related genes are transcribed as a single unit, called an "operon" (e.g.,
the lac operon, see below).
 • Translation: protein synthesis from mRNA. In prokaryotes, translation and
transcription occur concurrently, while in eukaryotes these two processes occur in
separate sub-cellular locations and sequentially in time.

• The genetic code: triplets of nucleotides; a total of 64 such triplets (codons) code
for a total of 20 kinds of amino acids (one type of amino acid can therefore be
coded by more than one codons); ATG is the start codon (coding for the beginning
of a protein) and there are 3 stop codons (signaling the end of translation).

Bacterial Gene Regulation

• Gene regulation: Protein synthesis is an energy-costly process. Therefore, many
bacterial genes are expressed (i.e. being transcribed and translated into its
product) only when needed. Regulation of gene expression (meaning the
switching-on and -off of genes) is an energy-saving strategy for a bacterial cell.

• Feedback inhibition at enzyme-pathway level: Inhibition of threonine (an amino

acid) synthesis pathway due to the binding of end product to the first enzyme of
the pathway. Feedback inhibition prevents cell from over-producing threonine.

• "Operon" refers to a group of genes and its regulators (e.g. a promotor) arranged
linearly as a unit of gene regulation. Operon regulation is at the DNA transcription
level (see two examples below).

• The lac operon: lactose is an uncommon sugar so genes for metabolizing lactose
is normally turned off (the promoter is blocked by a protein called repressor).
When lactose becomes available, it binds to the repressor and removes the
repressor from its DNA-binding site, resulting in the transcription of genes. As
the result, lactose is metabolized into carbon and energy source for the cell. This
turning-on of lactose-utilizing genes by the presence of lactose is an example of
enzyme induction (i.e., switching-on of genes when a chemical is present).

• The trp operon: tryptophan is an essential amino acid and expensive to

synthesize. Therefore, when enough tryptophan has been produced in a cell,
further synthesis of it is stopped. This turning-off of trptophan-synthesizing genes
by the pathway product itself is an example of enzyme repression (i.e.,
switching-off of genes when a chemical is present).

Mutation
• Mutation: changes in the genetic material (i.e. DNA) of a cell
• Type of mutation:
o Point mutation: change of a single base pair on a DNA molecule.
 Missense (Nonsynonymous) mutation: turning a codon coding for
amino acid A to a codon coding for amino acid B
  Nonsense mutation: turning a codon coding for amino acid A to a
stop codon
 Synonymous mutation: the original codon and the mutated codon
code for the same amino acid. No change of amino acid
 Transition (A<->G, C<->T) vs Transversion (Pyrimidine <-
>Purine)

o Insertion/Deletion (Indels): deletion of insertion of an extra base disrupts

three-by-three grouping of codons. This causes frameshift, introduces
premature stop codons, and results in truncated defective proteins.
• Mutagens (DNA-damaging agents): Chemical mutagens: nitrous acid, nucleotide
analog (used as anti-viral drugs, e.g. AZT). Radiation: UV light produces
thymine dimers between 2 neighboring T's on a DNA strand; X-ray and gamma
rays break DNA.
• Two experiments proving that adaptive mutation occurs randomly and is not a
result of exposure to antibiotics
o Fluctuation Test: If mutation occurs only when bacteria are exposed to
streptomycin, no difference in colony number fluctuation should be seen,
between the bacteria growing in a large or in small batches.
o Replica Plate Test: Antibiotic-resistent clonies on the master plate occur
without exposure to antibiotics.
• Ames test for chemical carcinogens: reversion of his- (histidine-dependent)
strains of Salmonella to his+ (histidine-utilizing) indicates a chemical being
mutagenic (therefore likely to be carcinogenic as well)