Microbiology- Chapter 8 Bacterial Genetics

If a base substitution occurs in a protein-coding region it can lead to what?

Silent or synonymous mutation - change in nucleotide leads to a codon that translates into the correct amino acid.

HW QUESTION- In transformation, genes from a donor chromosome are transferred as

Single-stranded DNA

Adding or deleting nucleotides during DNA replication is a...

Spontaneous mutation This can cause the loss or addition of an amino acid.

Competence for transformation

The recipient cell must be competent - physiological state that allows them to take up DNA Most bacteria take up DNA regardless of origin but some only accept DNA only from a closely related bacteria Some species are always competent but others need certain conditions.

Transduction

Transduction: transfer of bacterial genes by bacteriophages (phages) Phages are viruses that only infect bacterial cells The phage attaches itself to a bacterial cell and injects its nucleic acid (DNA - green) Phage enzymes cut bacterial DNA into small pieces (destroys the bacterial chromosome - purple) The phage takes over the bacterial cell's enzymes, which produce phage nucleic acid and a phage coat - components of new phage particles Phage particles are released from a bacterial cell, which is destroyed A transducing particle could be produced, phage picks up a piece of the bacterial chromosome instead of phage DNA (pink phage with purple instead of green)

HW QUESTION- Which of the following requires that cells be competent?

Transformation

Auxotroph

a mutation that requires that a growth factor be supplied to bacteria to survive. Scientist can remove the gene that codes for tryptophan (amino acid) synthesis. Organism can only grow if tryptophan is in growth medium (broth or agar). This allows scientists to keep track of and work with specific organism types.

Transposons

are introduced to cells to induce mutations They insert themselves into the bacterial genome, inactivating it, Transposons do not have an origin of replication Cannot be replicated!!!

Chemicals can change...

base-pairing properties Increases the chance that an incorrect nucleotide will be incorporated by DNA polymerase during DNA replication

Spontaneous mutations

changes that occur to genetic material from normal cell processes. Not a common occurrence As DNA replicates during cell division, mutations chances are between 1 : 10,000 and 1 : 1 trillion

Induced mutations

changes to the genome (DNA sequence) that are caused by outside influences Caused by a mutagen

Mutation

changes to the nucleotide sequence of DNA in cell. -Mutations are passed onto to progeny (daughter cells) -This is referred to as vertical gene transfer!

Prototroph

does not require any growth factors. This would be a bacterial cell that is found in nature (wild type)

Proofreading

done by DNA polymerases by checking for accuracy Can back up (reverse) and remove the incorrect nucleotide Inserts the correct nucleotide Very efficient but not perfect...

Mixmatch repair

fixes errors missed by DNA polymerase Enzyme cuts sugar-phosphate backbone of new DNA strand Another enzyme degrades short region of DNA strand with error DNA polymerase fills in the gap and DNA ligase seals it

HW QUESTION- A culture of E. coli is irradiated with UV light. The UV light affects the cell's DNA by

forming covalent bonds between thymine bases on the same strand of DNA.

Base analogs

have a nucleobase structure but do not hydrogen bond the same way Analogs are mistaken for nucleobases by DNA polymerase and used to synthesize DNA Analogs base pair with the wrong nucleobases

Intercalculating agents

increases the chance of frameshift mutations occurring They intercalate by laying flat and going in-between base pairs This pushes base pairs apart Base pairs can be deleted Ex. Ethidium bromide (used to visualize DNA separated during gel electrophoresis)

Transposons can cause...

insertional inactivation - inserted gene disrupts the function of a gene

Transformation

involves uptake of "naked DNA" Naked DNA is DNA not within a cell or virus Originates from cells that have been lysed (burst open) or secreted DNA

Genotype

is the sequence of nucleotides (ATGC) in DNA. This is changed when DNA is altered/mutated. There is only one set of genes in bacteria (haploid), so a small change can have a major impact.

SOS repair

last-ditch repair mechanism used when other systems have failed Induced following extensive DNA damage that stalls DNA and RNA polymerases Several dozen genes in SOS system Includes a DNA polymerase that synthesizes even in extensively damaged regions Has no proofreading ability, so errors are made Results in SOS mutagenesis The cell may survive with mutations or die from the damage

Bacteria can change their genetic material in what two ways?

mutation or horizontal gene transfer.

Phenotype

observable, physical, characteristics of an organism (capsule, flagella). Changes in the genotype alter these characteristics.

Base substitution

occurs during DNA synthesis when an incorrect nucleotide is added (incorrect complimentary base pairing).

Nonsense mutation

occurs when incorrect nucleotide substitution leads to the creation of a stop codon. This creates a shorter and most likely nonfunctioning protein. Called a knockout mutation.

Point substitution

one base pair changes. Can cause changes to a protein sequence (DNA to RNA to Protein).

Transposon

pieces of DNA that can move from one location to another in a genome Jumping genes move during transposition.

Frameshift mutation

the addition or deletion of one or two nucleotide pairs This changes the reading frame of transcribed mRNA and create a completely different set of codons

Horizontal gene transfer

the movement of DNA from one organism to another. These changes will be passed onto the progeny through vertical transfer.

Exposing cells to ultraviolet (UV) light causes the formation of...

thymine-thymine dimers Covalent bonds form between adjacent (neighboring) thymine DNA cannot fit into a double helix. Replication and transcription stop and the cell dies.

Missense mutation

when the wrong nucleotide is added it results in an altered codon that codes for a different amino acid. The effect of this mutation depends on the position and nature of the change within the protein.

HW QUESTION- Repair mechanisms that occur during DNA synthesis are

1. mismatch repair 2. proofreading by DNA polymerase

HW QUESTION- In which of the following does the donor cell direct the process?

Conjugation

Plasmid transfer

Conjugative plasmids direct their own transfer F plasmid (F stands for fertility) of E. coli most studied Encodes F pilus (sex pilus) F+ cells contain the F plasmid and F- cells do not

Conjugation

DNA transfer between bacterial cells Requires contact between donor and recipient cells

HW QUESTION- Which of the following types of repair does not require the standard DNA polymerase with proofreading capability?

SOS repair

The process of transformation

Double strand of DNA binds to cell surface and single strand of DNA enters the cell New DNA integrates into chromosome by homologous recombination Happens if the donor DNA is similar to the recipients Donor DNA replaces a homologous segment, which gets removed and degraded One daughter cell will inherit donor DNA Transformed cells can be detected by growing on medium that will not allow growth of non-transformed cells

Repair of errors in nucleotide incorporation

During replication, DNA polymerase sometimes incorporates the wrong nucleotide Mutation is prevented by repairing before DNA replication Two mechanisms of repair: Proofreading and Mismatch repair

Repair of Damaged DNA

Enormous amount of spontaneous and mutagen-induced mutations damage DNA If not repaired, can lead to cell death (in single-celled organisms) or cancer in animals Mutations are rare because alterations in DNA generally repaired before being passed to progeny

Example of chemicals that modify nucleobases.

Ex. Alkalizing agents add alkyl groups (short carbon chains) onto nucleobases Guanine is altered it base pairs to Thymine (should be Cytosine!) Turns to Adenine after another round of DNA replication

Types of Conjugation: Plasmid transfer steps

F pilus binds to a receptor on recipient cell wall F pilus contracts, pulling cells together Enzyme cuts plasmid at origin of transfer Single DNA strand is transferred Complementary strands synthesized Both cells are now F+

Types of Conjugation: Chromosome transfer

F plasmid is integrated into a chromosome by homologous recombination Involves Hfr cells (high frequency of recombination) Process is reversible F′ plasmid results when small piece of chromosome is removed with the integrated F plasmid DNA F′ plasmid is transferred to F- cells Carries bacterial DNA into new cells Hfr cell produces the F pilus Integrated F plasmid begins the transfer of genes on one side of origin of transfer of plasmid Part of the chromosome is transferred to recipient cell, chromosome usually breaks before complete transfer Recipient cell remains F− since incomplete F plasmid transferred

Generalized transduction

Generalized transduction results when a fragment of bacterial DNA enters the phage protein coat Produces a transducing particle Transducing particle may attach to another bacterial cell and inject the DNA it contains New DNA may be integrated into chromosome

HW QUESTION- Match the chemical mutagens with their descriptions.

Intercalating agents- insert between base pairs. Base analogs- used in place of normal nucleobases. Transposons- randomly insert into DNA. Alkylating agents- add CH3 and others to nucleobases.

Repair of Modified Nucleobases in DNA

Modified nucleobases lead to base substitutions (incorrect base pairing) The enzyme Glycosylase removes the oxidized nucleobase (base substitution) Another enzyme cuts DNA at this site DNA polymerase removes short section and synthesizes its replacement DNA ligase seals the gap

HW QUESTIONS- Please match each type of mutation with its appropriate description.

Nonsense -A mutation that changes a codon that specifies an amino acid to a stop codon, resulting in premature termination of polypeptide synthesis. Missense- A mutation that results in changing a codon such that a different amino acid is specified. Silent mutation- A mutation that changes a codon into a different codon, but both codons specify the same amino acid; this causes no change in the resulting polypeptide. Frameshift- A mutation resulting from an insertion or deletion of bases that causes a change in the reading frame of the mRNA.

Repair of Thymine dimers

Photoreactivation: light repair Enzyme uses energy from light Breaks covalent bonds of thymine dimer Only found in bacteria!!! Excision repair: dark repair Enzyme removes damage DNA polymerase fills in the gap DNA ligase seals it