Chapter 8: Bacterial Genetics

mobile gene pool

-Genomics reveals surprising variation in gene pool of even a single species -perhaps 75% of E. coli genes found in all strains, termed *core genome* of species. -remaining make up the _______ : plasmids, transposons, genomic islands, phage DNA.

Plasmid

A DNA molecule that replicates independently of the chromosome (element of the mobile gene pool) -found in many Bacteria and Archaea; some Eukarya -usually circular double stranded DNA molecules, with origin of replication -generally encodes nonessential information; cells survive loss -important because they provide cells with the ability to survive in a particular environment -vary with respect to their properties

Mismatch repair

A process that fixes errors missed by the proofreading of DNA polymerase in which a specific protein binds to the site of the mismatched nucleobase, directing an enzyme to cut the sugar-phosphate backbone of the new DNA strand. Another enzyme then degrades a short region of that DNA strand, removing the misincorporated nucleotide.

Mutation

A change in the nucleotide sequence of a cell's DNA that is passed on to daughter cells

auxotroph (auxo= increase/ troph=nourishment)

A microorganism that requires an organic growth factor

SOS mutagenesis

A process in which errors are made due to the lack of proofreading ability in SOS repair of extensively damaged DNA

Proofreading

DNA polymerases are complex enzymes that not only synthesize DNA, but also check the accuracy of their actions- this process is called: The enzymes can back up and remove a nucleotide not correctly hydrogen-bonded to the opposing nucleobase in the template strand & inserts the correct nucleotide

non-homologous recombination

DNA recombination that does not require extensive nucleotide sequence similarity in the stretches that recombine

horizontal gene transfer

DNA transfer from one bacterium to another by conjugation, DNA-mediated transformation, or transduction

Specialized transduction

During this, in the lysogenic cycle, the prophage eventually excises itself from the host chromosome. Sometimes it takes a few nearby host genes and leaves a few phage genes behind

thymine dimers

Exposing cells to UV light causes covalent bonds to form between adjacent thymine nucleobases on a DNA strand, producing ______, which distort the DNA molecule because they cannot fit properly into the double helix. Replication and transcription stop at the distortion, and cells will die as a result if the damage is not repaired (Mutations result from cell's SOS repair mechanism)

Replicon

Following gene transfer, recipient cells must replicate the DNA to pass it on to daughter cells. This can happen only if the DNA is a ______, meaning it has an origin of replication. (Ex: plasmids and chromosomes; NOT fragments of chromosomal DNA)

F' (F prime) donors

Hfr strains can revert to F+ because the process of F plasmid integration is reversible. In some instances, an error can occur during excision, and a piece of the bacterial chromosome is removed along with the F plasmid DNA. This action brings a chromosomal fragment into the F plasmid, producing a plasmid called ______. Like the F plasmid, this is a replicon that is rapidly and efficiently transferred to F- cells. However, in this case, the chromosomal fragment is transferred as well.

point mutation

If only ONE base pair is changed it is a:

Photoreactivation (light repair)

In a repair mechanism found only in microbes (bacteria), an enzyme uses the energy of visible light to break the covalent bonds of the thymine dimer, restoring the DNA to its original state. Because light is required for this mechanism, it is called:

natural selection

In the ever changing conditions that characterize most environments, organisms need to adapt in order to survive and multiply. If they fail to do this, competing organisms more fit to thrive in the new environment will soon dominate. This is the process of:

Methylation of the DNA nucleobases

In the process of mismatch repair, how does the cell know which strand is the new one?

Lysogenic cycle

In this cycle of transduction, temperate phages integrate their DNA into the host chromosome (as a prophage)

Lytic cycle

In this cycle of transduction, virulent phages: -destroy the host chromosome -replicate themselves & -destroy the cell.

silent or synonymous mutation

In this type of mutation, the change generates a codon that translates into the same amino acid as the original. This can occur because of the redundancy of the genetic code "Wild-type amino acid"

Genomic island

Large genomic segment in a cell's genome that originated in another species (element of the mobile gene pool)

Carcinogens ("cancer-generating")

Many cancers appear to be caused by chemicals that are ______, and most are mutations. Animal tests for screening are expensive & time consuming.

restriction enzymes

Recognizes a specific short nucleotide sequence within the cell and then cuts the DNA molecule at that sequence

Barbara McClintock

Scientist who studied color variation in corn kernels & concluded that segments of DNA (transposons or "jumping genes") we're moving into and out of genes involved with kernel color. These moving DNA segments destroyed the function of the genes, thereby changing kernel color. Received Nobel Prize in 1983 for this discovery

penicillin enrichment

Selectively kills prototrophs and is sometimes used before replica plating to increase the proportion of auxotrophic mutants, which makes the cells easier to isolate. Relies on the fact that penicillin kills only growing cells. The cells that have been treated with the mutagen are incubated in glucose-salts broth containing penicillin

Insertion sequence (IS)

Simplest type of transposon which encodes only transposase (the enzyme responsible for transposition) *on each side of the gene are inverted repeats- sequences that are identical when read in the 5' to 3' direction

Spontaneous and mutagen-induced

The amount of _____ and _____ damage to DNA is enormous, and if it is not repaired, can quickly lead to cell death (leads to cancer in animals) For example: in humans, two genes associated with breast cancer code for DNA repair enzymes; mutations in either result in 80% probability of breast cancer

pathogenicity islands

The characteristics encoded by genomic islands include use of specific energy sources, acid tolerance, development of symbiosis, and ability to cause disease. Genomic islands that encode the ability to cause disease are:

transducing particle

The error resulting from generalized transduction creates a ____, which carries no phage DNA and therefore is NOT a phage, but will ONLY attach to another bacterial cell and inject the DNA it contains

Insertional activation

The gene into which a transposon jumps is inactivated by the event, an outcome called ______, meaning that the inserted DNA disrupts the function of the gene.

base substitution

The most common type of mutation; occurs during DNA synthesis when an incorrect nucleotide is incorporated

Phenotype

The observable characteristics of a cell

Genotype

The sequence of nucleotide in an organisms DNA

Gene pool

The sum of all genes

X-rays

These cause single- and double-strand breaks in DNA, and changes to the nucleobases. Double-strand breaks often result in deletions that are lethal to the cell

intercalating agents

These increase the frequency of frameshift mutations. They do this because they are flat molecules that can insert between adjacent base pairs in a strand of DNA. This pushes the nucleotides apart, producing a space between bases that allows errors to be made during replication

base analogs

These structurally resemble nucleobases but have different hydrogen-bonding properties. These can be mistakenly used in place of the nucleobases when the cells make nucleotides, and DNA polymerase then incorporates these into DNA. When the complementary strand is synthesized, the wrong nucleotide may be incorporated opposite of this..

restriction modification systems

These were discovered by scientists studying why certain bacterial strains are relatively resistant to phage infection. Their research showed that those strains degrade foreign DNA through the combined action of two types of enzymes: a restriction enzyme and a modification enzyme.

Generalized transduction

results from a rare error that sometimes occurs during the construction of phage particles. A fragment of bacterial DNA- produced when the phage-encoded enzyme cuts the bacterial genome- mistakenly enters the phage protein coat during packaging, which they transfer to a new host cell

Transposon

segment of DNA that can move from one site to another in a cell's genome; directs its own movement to another location in chromosomal or plasmid DNA (element of the mobile gene pool)

mutation rate

the probability that a mutation will occur in a given gene per cell division

vertical gene transfer

transfer of genes from an organism to its offspring

Replica plating

A process used to overcome the problems that arise from indirect selection of auxotrophic mutants. Step 1: the master plate of bacterial colonies is pressed onto the surface of a sterile velvet, which picks up cells from every colony. Step 2: two agar plates(one nutrient agar and one glucose-salts agar) are pressed in succession and in the same orientation onto the same velvet. Cells adhering to the velvet are transferred to the sterile media, resulting in exact replicas of the original plate. Step 3: replica plates are incubated, allowing auxotrophic mutants to form colonies on the nutrient agar but not on the glucose-salts agar

Excision repair (dark repair)

A repair mechanism in which an enzyme recognizes the major distortions in DNA that result from thymine dimer formation and then removes the DNA strand with the damaged region. DNA polymerase and ligase then fill in and seal the gap left by removal of the segment.

SOS repair (last-ditch repair)

A repair mechanism that is a last-effort attempt that bacteria use to repair extensively damaged DNA. The enzymes that carry out this repair are induced when photoreactivation and excision repair are unable to correct damages. DNA and RNA polymerases stall at sites of unrepaired damage. Several dozen genes are activated (including a DNA polymerase that synthesizes even in extensively damaged regions) Has NO proofreading ability, so errors are made *without this cells would die*

Competence

A specific physiological state that allows a recipient cell to take up DNA from the environment, in order for transformation to occur. -most competent bacteria take up DNA regardless of its origin (some accept only from closely related bacteria, recognizing it by DNA sequence) -it may give the recipient the ability to be more pathogenic

The Ames Test

A test that uses a histidine-requiring auxotroph of Salmonella typhimurium (His-) to measure the mutagenicity of a chemical. This agar medium used for this type of testing lacks histidine, so His- auxotrophs cannot grow, whereas prototrophs can. -uses direct selection -if mutagenic, reversion rate increases relative to control -rat liver extract may also be added since non-carcinogenic chemicals are often converted to carcinogens by animal enzymes -if the test chemical causes mutation of His- cells, additional testing must be done in animals to determine if it is a carcinogen (although most carcinogens are mutagens, not all mutagens are carcinogens)

sex pilus/pili

A tube between (+) and (-) prokaryote cells in which the cell can send copies of its plasmids that can contain genes for things like: the ability to make sex pili (which would change the cell from (-) to (+), also possible to contain genes for things such as antibiotic resistance)

frameshift mutation

Adding or subtracting one or two nucleotide pairs causes a _____. This changes the reading frame of the corresponding mRNA molecule so that an entirely different set of codons is translated

Mutagens

An agent that induces change is a ______. Geneticists may use these to increase mutation rate **TWO general types: chemical and radiation**

"null" or "knockout mutation"

Any mutation that totally inactivates the gene is termed a

Aerobic/Anaerobic

Base substances are more common when bacteria are growing in ______ environments, as opposed to ______ ones

Hfr cells (high frequency of recombination cells)

Chromosomal DNA transfer is less common than plasmid transfer and involves ______. These are strains in which the F plasmid has integrated into the chromosome by homologous recombination, which happens on rare occasions.

DNA-mediated transformation

Mechanism of horizontal gene transfer in which "naked" DNA is taken up by the recipient cell from its surroundings. (Naked means there are no coverings like a membrane or a nucleus around the DNA) this is very commonly carried out in bacterial labs where DNA is heat shocked, which loosens membranes and allows the cells to absorb DNA more easily)

Conjugation

Mechanism of horizontal gene transfer in which the donor cell physically contacts the recipient cell usually transferring a plasmid (this mechanism is the closest that prokaryotes have gotten to sexual reproduction) *most commonly studied example is the F plasmid (F stands for fertility) F (+) cell has the ability to make a mating bridge called a sex pilus with another bacteria that CANNOT make sex pili F (-) {Gram positive and gram negative can both transfer DNA this way but the process is different between the two.}

Recombinants

Micro organisms commonly acquire genes from other cells, through a process called horizontal gene transfer. To determine if the recipients have indeed acquired new characteristics, the donor and recipient cells must be genetically different, which is one reason mutants are useful. The resulting cells called _____, have properties of each of the original strains.

Direct selection

Mutants that can grow under conditions in which the parent cells cannot are usually easy to isolate by ______. In this method, cells are inoculated onto an agar medium that supports the growth of the MUTANT, but does not allow the parent to grow. Ex: antibiotic resistant mutants can be easily selected directly by inoculating cells onto a medium containing the antibiotic; only the resistant cells will form colonies

Low-copy-number plasmids

Occur in only one or a few copies per cell

Reversion

On rare occasions a mutation will change back to its original non-mutated state. This change is called a ______, or back mutation, and like the original mutation, occurs spontaneously at low frequencies.

antibiotic resistance

Organisms in which some plasmid-encoded traits are found: -many including Escherichia coli, Salmonella species, staphylococcus species... Name the trait:

Tumor formation in plants

Organisms in which some plasmid-encoded traits are found: Agrobacterium species Name the trait:

toxin production

Organisms in which some plasmid-encoded traits are found: Bacillus anthracis Name the trait:

Insect toxin synthesis

Organisms in which some plasmid-encoded traits are found: Bacillus thuringiensis Name the trait:

pilus synthesis

Organisms in which some plasmid-encoded traits are found: E. Coli, Pseudomonas species Name the trait:

Gas vacuole production

Organisms in which some plasmid-encoded traits are found: Halobacterium species Name the trait:

Oil degradation

Organisms in which some plasmid-encoded traits are found: Pseudomonas species Name the trait:

nitrogen fixation

Organisms in which some plasmid-encoded traits are found: Rhizobium species Name the trait:

Antibiotic synthesis

Organisms in which some plasmid-encoded traits are found: Streptomyces species Name the trait:

Increased virulence

Organisms in which some plasmid-encoded traits are found: Yersinia and Shigella species Name the trait:

Phage DNA

Phage genome that sometimes carries additional genes (element of the mobile gene pool)

homologous recombination

Process by which a cell replaces a stretch of DNA with a segment that has a similar nucleotide sequence

modification enzymes

Protects that cell's own DNA from the action of the restriction enzyme by adding methyl groups to the nucleobases recognized by the restriction enzyme (which will destroy incoming foreign DNA but not the host DNA)

CRISPR Systems

These were discovered when scientists recognized that certain bacterial genomes include very small pieces of phage DNA that recognize the Specific DNA if it invades the cell again. -first invasion: complex of Cas proteins cuts DNA into short fragments -some of those DNA fragments are then inserted into a chromosomal region called a CRISPR array -an integrated segment of captured DNA is called a spacer

missense mutation

This type of mutation results when the altered codon codes for a different amino acid. The effect of this depends on the position and the nature of the change within the protein. In many cases, cells with this mutation grow slowly because the encoded protein does not function as well as normal... "leaky"

non-homologous recombination

Transposons integrate into their new location through this process that does not require a similar nucleotide sequence in the region of recombination. The transposon simply inserts a stretch of DNA; it does not replace the existing sequences

Ultraviolet (UV) light and X rays

Two kinds of radiation that are commonly used as mutagens are:

"Proofreading" by DNA polymerase & mismatch repair

Two mechanisms that repair damaged DNA before errors are replicated are: By quickly repairing the errors before replication, the cell prevents the mutation.

nonsense mutation

Type of mutation that occurs when the base substitution creates a stop codon, resulting in a SHORTER (truncated) and often non-functional protein.

Composition transposons

Type of transposon which consists of one of more genes flanked by ISs. Like insertion sequences, these can move in the same replicon or from one replicon to another in the cell. Their movement is easily followed if they encode a recognizable gene product such as antibiotic resistance.

Subsequent invasion

When transcription of CRISPR array generates crRNA's that direct DNA-cutting enzymes to invading DNA

Transposition

When transposons are introduced intentionally into a cell in order to generate mutations it is called ____. In this process, the transposon, which can not replicate on its own because it lacks an origin of replication, inserts into the cell's genome, which generally inactivates the gene into which it inserts

Resistance or R plasmids

____ are particularly important medically because they encode resistance to antimicrobial chemicals, including antibiotics. Many are conjugative, carrying out only genes encoding resistance (R genes) but also genes for pilus synthesis and other properties required for conjugation. Members of the normal microbiota that carry these cans potentially transfer them to pathogens

Indirect selection

_____ is used to isolate an auxotrophic mutant from a prototrophic parent strain. This process is more difficult because any medium that supports the growth of the mutant also allows the growth of the parent.

Mutations

______ are rare because cells have multiple mechanisms to repair damaged DNA before errors are passed on to progeny. However, these mechanisms are not perfect and certain types of DNA damage such as insertional inactivation caused by transposons cannot be repaired.

Conjugative plasmids

______ direct their own transfer from donor to recipient cells.

Modified nucleobases

____________ can result in base substitutions if they are not repaired before the DNA is replicated. An example of a repair mechanism for this involves oxidized guanine

prototroph (proto=earliest form of/troph=nourishment)

a microorganism that does not require any organic growth factors

induced mutations

genetic changes that occur due to an influence outside of a cell, such as exposure to a chemical or radiation

spontaneous mutations

genetic changes that result from normal cell processes; they occur randomly and happen at infrequent but characteristic rates

Transduction

mechanism of horizontal gene transfer in which bacterial DNA is transferred using viruses inside a phage coat. A bacteriophage carries a chromosomal DNA fragment from donor to recipient.