portage learning genetics- mod 4

bacteriophages

viruses that use bacteria as a host

why are bacteria great tools for genetic research?

- they grow quickly; have a short reproductive cycle
- they are easy to isolate 1 type of bacteria using antibiotic resistant strains
- they can be grown in almost any lab with little risk
- their nourishment is inexpensive

what is a defining characteristic of bacteria?

spontaneous mutation

what is spontaneous mutation

the process by which bacteria undergo random mutations
- bacteria developed a resistance to T1 infection through mutation aka, adaptive immunity

who presented an experiment showing that bacteria are capable of random spontaneous mutation in the 1940s?

salvador luria & max delbruck

what is selection?

it involves choosing bacterial colonies in conditions where the desired mutation grows, but the wild-type doesn’t

how do scientists use selection?

by adding a resistance marker to a dna plasmid that contains genes of interest so they can grow large amounts of bacteria that contain the desired gene

what are the 2 ways bacteria are grown in labs?

- in liquid culture

- on agar (semi-solid gelatinous material)

what are bacteria usually grown in?

minimal medium

what are minimal medium?

a growth medium where bacteria can synthesize all necessary organic compounds (salts, sugars, etc) for sustenance

prototrophic

bacteria that can grow on minimal media and don’t need additional nutrients or reagents to grow

auxotrophic

bacteria that lack essential nutrients or amino acids and can’t grow without supplementation

what are the phases of bacterial growth?

lag phase, log phase, stationary phase

lag phase

growth is very slow

logarithmic (log) phase

a fixed amount of time between periods of replication resulting in exponential growth

stationary phase

bacteria ceases to grow

when is the best time for scientists to harvest bacterial culture?

during the log phase of growth to ensure maximum health

what are the ways bacteria can transfer genetic information to another bacteria by?

conjugation, transformation, transduction

what is conjugation?

the process by which one bacterium transfers genetic material to another bacterium via direct contact

what did the experiment joshua lederberg and edward tatum did to demonstrate conjugation conclude?

spontaneous mutation restoring all function was unlikely and genetic recombination between strains had occurred

F+ cells

donor bacteria that donate genes
- F is for fertility

F- cells

recipient of the genes

- these cells receive genetic info from donor and integrate it into their own chromosomes

what is required for conjugation

physical cell to cell contact

F-pilus

a structure that mediates contact with the cell for conjugation

Fertility factor (f-factor)

aka plasmid, circular double stranded dna

- cells that contain this can donate genetic material

plasmids

- include genes needed for transfer of genetic material
- only cells with plasmids can form an f-pilus and transfer dna to f- cells

what is one function of plasmids?

giving bacteria resistance to multiple antibiotics

where are plasmids located?

within cytoplasm of bacterial cell

how do plasmids replicate?

by using the host cell’s enzymes just like bacterial chromosomes

how do plasmids exist?

in high- copy or low-copy numbers and are passed along with the host chromosome to daughter cells

high copy/ low copy

produce high expression of plasmid, while the opposite is true about low-copy plasmids

R- plasmids

bacteria with antibiotic resistance plasmids

what are the 2 key components of r plasmids?

- RTF ( resistance transfer factor)

- R-determinants

-RTF ( resistance transfer factor)

enables transfer of genetic material between bacteria
- RTFs are genetically similar across different bacterias

r-determinants

encode specific antibiotic resistance genes

what are some common resistances to antibiotics

ampicillin, tetracycline, kanamycin, chloramphenicol, and streptomycin

what are some harmful things about bacterial R plasmids?

they confer pathogenic resistance in the medical community, rendering dangerous bacteria impervious to a wide variety of antibiotics
(i.e MRSA- a staph bacterial infection)

what are some useful things about bacterial R plasmids

they are valuable tools in genetic research

what is transformation?

the result of a bacterium taking up extracellular dna from its environment and incorporating it stably into its genetic makeup

what is the criteria for bacteria to be transformed?

it must be physiologically competent

How is bacteria induced to competency if it already isn’t?

through the use of chemicals in conjunction with heat or electroporation methods (E.coli)

how do competent bacteria take up dna in transformation process?

by opening small pores in their membranes that allow direct passage of dna

what happens after dna enters the cell in transformation process?

dna is digested by nucleases (enzyme), leaving one single strand of dna, which can can align in the complementary region of the bacterial chromosome and replace its counterpart

what is the dna in the bacteria known as after it replaces the original dna?

heteroduplex

what is a heteroduplex?

dna with 2 sources of nucleotides that are not identical matches

what happens in the last step during the process of transformation?

dna repair mechanisms are initiated and after one round of dna replication, a transformed (mutant) cell and untransformed (original) cell are produced

phages

aka bacteriophages, are viruses that use bacteria as their host and transmit their genetic material via transduction

what is the life cycle of a phage?

1. base plate of phage binds to surface of cell

2. phage dna from outer sheath is injected through cell membrane & host cell dna is degraded

3. phage dna is replicated & phage proteins are synthesized

4. mature phages are assembled

5. bacterial membrane is lysed as phage is released from cell

what are 3 types of phages?

- virulent phages

- prophages

- temperate phages

virulent phage

viruses that always lyse their bacterial host

prophages

viral dna that integrate into bacterial chromosome

- viral and bacterial genetic material are produced in a way that doesn’t kill bacteria

- aren’t replicated til a stimulation event

temperate phages

viruses that are capable of lysis or that act as a prophage

- intracellular conditions determines how the virus will act

what is transduction?

process of bacterial genetic material being transmitted by viruses with the absence of contact

what are some things that caused drug resistance to antibiotics?

The fact that 80% of the world’s antibiotics are given to livestock and antibiotics used to be given to treat and and every illness even if it wasn’t bacterial

what are some problems with making new antibiotics?

- it is difficult to make antibiotics because most of them are broad spectrum, which kills bad and good bacteria, making them difficult to develop as drugs

- they are not money makers for producers, so they’re not at the forefront of being frequently produced

what are pros of phage therapy?

- they can be engineered to be bacterium specific

- it does not require long term therapy

what are cons of phage therapy?

- long term consequences are unknown

- phage resistance could arise

- doctors must be 100% certain as to what the bacterial source is

cotransduction

genes that transduce simultaneously via phage

basal level

most bacterial gene products are constantly produced in a few copies per cell, but when called upon, their activity can increase drastically; baseline

what is the cellular decision to moderate gene expression largely regulated by?

extracellular environment

- When conditions are ideal for cell growth and replication, the genes responsible for metabolic activity and growth will be activated

- when conditions are harsh, genes needed for cellular survival may activate

enzyme

a substance produced by a living organisms that acts as a catalyst to bring about a specific biochemical reaction

what are 3 types of enzymes

- inducible enzymes

- repressible enzymes

- constitutive enzymes

inducible enzymes

turned on under certain environmental conditions

- under positive control

- ex: lactose-metabolizing genes

repressible enzymes

turned off in response to certain conditions

- under negative control

- ex: tryptophan synthesis genes; when it’s available from the environment, bacteria won’t expend energy on tryptophan synthesis

constitutive enzymes

produced all the time, irrespective of the environment or cellular conditions

positive control

gene expression is turned off until regulator enzymes activate them

negative control

gene expression continues until told to shut off by a regulator molecule

operon

cluster of genes that are typically located together

lac operon

functions to turn the metabolic activity of lactose on or off

structural genes

not involved in regulation but form the structure of an enzyme

what are the 3 structural genes of the lac operon

- lacZ gene

- lacY gene
- lacA gene

lacZ gene

this gene encodes for for b-Galactosidase, which converts lactose (a disaccharide) into glucose and galactose

lacY gene

this gene encodes for permease, an enzyme that facilitates lactose entry into the bacterial cell

lacA

lacA gene encodes for the enzyme transacetylase, which is thought to help to remove toxins that are built-up by lactose digestion

regulatory region

a structure that lies just upstream from the three structural genes

what do studies with lac operon rely on?

chemical analogs of lactose such as isopropylthiogalactoside (IPTG), which is a gratuitous inducer

gratuitous inducer

they can activate genes naturally, but they do not act as substrates for the enzymes that are synthesized

lacl

makes repressor proteins when lactose isn’t around

how does the lacl gene act when lactose isn’t present?

the lacl gene produces the repressor, which binds to the operator region of dna and blocks genes needed to break down lactose

how does the lacl gene act when lactose is present?

the lacl gene binds to the repressor and changes its shape, which causes the repressor to let go dna, allowing lactose metabolizing genes to turn on

what do sRNAs do in bacteria?

they help regulate gene expression by binding to gene sequences to block or enhance gene activity

- they are short(50-100 nucleotides)

what do sRNAs do during stressful conditions (cold or environment changes)?

they help bacteria in a stationary phase survive by turning on survival genes (like hibernation)

what is an example of what sRNA does?

- RyhB sRNA in E. coli is a negative regulator of genes that control iron expression

- when iron levels are low, RyhB will inhibit nonessential genes that require iron, which allows critical enzymes to use the limited supply of iron