question

The central dogma

answer

DNA --> RNA --> Protein
Replication --> Transcription --> Translation

question

Multiple genes that are organized together on a genome:

answer

Operon

question

Operon

answer

A group of related genes arranged in a linear fashion whose expression is under control of a single operon --> allows for expression of multiple genes to be coordinated.

question

The backbone of the DNA chain

answer

alternating phosphates and the pentose sugar deoxyribose

question

What eukaryotic cell has plasmids?

answer

Yeast

question

Transposable elements

answer

segment of DNA that can move from one site to another site on the same or different DNA molecule.
Inserted into other DNA molecules.
three main types:
- insertion sequences
- transposons
- special viruses

question

Plasmids

answer

typically double stranded genetic elements, typically circular, that replicate separately from the chromosome.
may confer a selective growth advantage under certain conditions.
virulence characteristics are encoded by plasmid genes.
Genes encoding Bacterions are often carried on plasmids.

question

Episomes. Who are similar?

answer

plasmids that can integrate into the cell chromosome. Similar situation can be seen with prophages.

question

Word for the removal of plasmids from host cells.

answer

curing

question

What are the different kinds of plasmids?

answer

episomes (integrate into cell chromosome)
Conjugative (transfer via cell to cell contact)
R-plasmids - Resistant plasmids; confer resistance to antibiotics and other growth inhibitors

question

R-plasmids

answer

Plasmids that confer a resistance to antibiotics and other growth inhibitors.
Many on conjugative.

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Virulence factors

answer

enable pathogen to colonize
enable pathogen to cause host damage
hemolysin
enterotoxin

question

Bacteriocins

answer

Proteins produced by bacteria that inhibit or kill closely related species or different strains of the same species.

question

What are the different kinds of transposable elements?

answer

Insertion sequences (IS)
Transposons
Special viruses

question

Bacteriophages

answer

Viruses that take over a bacterial cell and force it to replicate the viral DNA.

question

Replication always proceeds from which end?

answer

5' to 3' ends.

question

Why is DNA said to be semiconservative?

answer

Because each of the two progeny double helices have one parental strand and one new strand.

question

What is the precurser to each nucleotide?

answer

a deoxynucleoside 5'-triphosphate (dNTP)

question

Why does replication proceed from 5' to 3'?

answer

in order to create new peptide bonds, a hydrolysis reaction must occur. This means that there must be a hydroxyl group available and this is only available on the 3' side.

question

DNA polymerases

answer

DNA polymerases catalyze the addition of dNTP's (deoxynucleoside 5'-triphosphates...the precursers toe ach nucleotide)

question

What do DNA polymerases require before they can begin?

answer

an RNA primer

question

what creates the RNA primer needed by DNA polymerase?

answer

RNA primase.

question

replication fork

answer

zone of unwound DNA where replication occurs.

question

DNA helicase

answer

an enzyme that unwinds the DNA for replication (etc)

question

What's the difference between the leading and lagging strands?

answer

Extension of DNA occurs continuously on the leading strand.
Extension of DNA occurs discontinuously on the lagging strand. These fragments are called Okazaki fragments.

question

Transcription is carried out by which polymerase?

answer

RNA polymerase

question

RNA polymerase

answer

carries out transcription of both DNA and RNA molecules.

question

What's the difference between RNA and DNA polymerases?

answer

RNA polymerase doesn't need a primer.

question

What does RNA polymerase recognize on DNA strands?

answer

sites that are called promoters. They are sites of initiation for transcription.

question

What part of the RNA polymerase recognizes the promoters?

answer

the Sigma factor (subunit)

question

what are the two regions within promoters that are highly conserved (and important) where are they located?

answer

Pribnow box (-10 region)
-35 region --> located 35 bases upstream of transcription.

question

Concensus sequences

answer

Sequences that establish a baseline rate of transcription for that gene. Controlled by how deviant from the ideal concensus sequence it is.
Found at the -10 and -35 base regions of the promoter sequence.

question

What terminates transcription?

answer

Termination of RNA synthesis is governed by a specific DNA sequence.
Intrinsic terminators: Transcription is terminated without any additional factors.
Rho-dependent termination: Rho protein recognizes specific DNA sequences (rho sequence) and cause a pause in RNA polymerase.
Inverted repeats will terminate transcription: A stem loop in RNA immediately upstream from a run of uracils leads to transcription termination.

question

Unit of transcription:

answer

Unit of chromosome bounded by sites where transcription of DNA to RNA is initiated and terminated.
Example: Operon or Regulon.

question

An RNA encoding a group of cotranscribed genes is called what?

answer

polycistronic mRNA

question

Regulon

answer

A group of operons that are coordinately regulated.

question

constitutive proteins

answer

proteins that are needed at the same level all the time.

question

What is the ribosome binding site?

answer

the Shine-Dalgarno sequence

question

What are the major modes of regulation?

answer

One controls the activity of preexisting enzymes
-posttranslational regulation (takes seconds)
One controls the amount of an enzyme
- regulate level of transcription
- regulate translation (minutes)

question

How is transcription regulated?

answer

DNA-binding protein,
Negative control of transcription: Repression and induction
Repression: protein binds to DNA in response to a signal and blocks synthesis of enzymes
Induction - A small molecule binds to a repressor and causes it to fall off, thereby synthesizing the enzyme.
Positive control of transcription: binding even of regulatory protein to DNA activates transcription
global control and the lac operon

question

DNA binding proteins

answer

small molecules that influence the binding of regulatory proteins to DNA. (proteins that regulate transcription).
Most DNA-binding proteins interact with DNA sequences with specificity.
Major groove of DNA is the main site of protein binding.

question

which groove of the DNA is the main site of protein binding?

answer

the major groove of DNA.

question

What type of sequence is typically the binding site for regulatory proteins?

answer

inverted repeats ==> read the same forwards as backwards and form a hairpin loop.
**remember terminators are also these

question

Homodimeric proteins

answer

Proteins composed of two identical polypeptides.
- Protein dimers interact with inverted repeats on DNA. Each of the polypeptides binds to one inverted repeat.

question

what are the classes of protein domains that are critical for proper binding of proteins to DNA.

answer

helix-turn-helix : First helix is the recognition helix. Second is the stabilizing helix.
Ex: Lac and trip repressors of e. coli

Zinc Finger : protein structure that binds a zinc ion.
There's typically 2-3 zinc fingers on proteins that use them for DNA binding.

Leucine Zipper : leucine residues are placed every seven amino acids
Does not directly interact with DNA, instead interacts through the hydrophobic effect.

question

Inducer

answer

a substance that induces enzyme synthesis by binding to a regulatory protein and causing it to fall off the DNA strand.

question

Corepressor

answer

a substance that represses enzyme synthesis. It binds to a regulatory protein, a repressor, and causes it to bind to DNA.

question

effectors

answer

collective term for inducers and repressors.
repressor molecules bind to an allosteric repressor protein.

question

Allosteric repressor

answer

Once bound to a repressor, allosteric repressors become active and bind to a region of DNA near the promoter called the operater.

question

where is the operater found? What is it?

answer

downstream from the promoter. It is the sequence where the regulatory proteins bind and block transcription.

question

Repressor

answer

A regulatory protein that, when bound to a corepressor, binds to a DNA strand and blocks transcription.

question

Positive control

answer

A regulator protein activates the binding of RNA polymerase to DNA.
Activator proteins, when bound to an inducer, bind to the activator binding site and RNA polymerase and urge transcription forwards.

question

Global control systems. what are some xamples of it?

answer

regulate expression of many different genes simultaneously.
Catabolite repression: synthesis of unrelated catabolic enzymes is repressed if glucose is present in the growth medium.
- Lac operon is under control of catabolite repression.

question

Diauxic growth

answer

Two exponential growth phases. Occurs when glucose is used up and another energy source has been tapped.

question

Catabolite repression is what kind of activation?

answer

positive activation

question

Selectable mutations

answer

Those that give the mutant a growth advantage under certain conditions

question

nonselectable mutations

answer

those that give the mutant neither an advantage or disadvantage over the parent. e.g. inability to grow under certain conditions. requires screening to detect mutants.

question

Bacteria that can grow on one medium but not another would be called what?

answer

auxotrophs

question

List and describe the different methods of mutation.

answer

induced mutations
- environmentally or deliberately induced
- exposure to oxygen radicals or natural radiation

spontaneous mutations

point mutations
- mutations that change only one base pair
- can lead to single amino acid change in a protein, an incomplete protein or no change at all (silent)

question

What are the different types of mutations that can occur?

answer

Silent mutation - does not effect amino acid sequence
missense mutation -AA changed - polypeptide altered
nonsense mutation - codon becomes stop codon, incomplete protein.
Frameshift mutations - deletions or insertions result in a shift of reading frame. Often results in complete loss of gene function.

question

Which type of mutation is usually reversable?

answer

point mutations.

question

reversion

answer

alteration of DNA that reverses the effects of a prior mutation

question

revertant

answer

stain in which the original phenotype has been restored.
two types:
same type revertant: mutation is at same site as original mutation
second-site revertant: mutation is at a different site in the DNA.

question

Mutagents:

answer

chemical, physical or biological agents that increase mutation rates

question

what are the classes of mutagents:

answer

chemical mutagens:
- nucleotide base analogs -> resemble nucleotides
- C.M. that induce chemical modification
- C.M. that cause frameshift mutations (example: acridine)

Radiation:
- non ionizing --> purines and prymidines strongly absorb UV. Pyrimidime dimer is an effect of this.
- Ionizing radiation (cosmic rays, xrays, gamma rays)High energy short wave. --> ionize water and cause free radicals which damage macromolecules in the cell

question

What are the three types of DNA repair systems?

answer

Direct Reversal - mutated base is still recognizable and can be repaired without referring to other strand.
Repair of single strand damage - damaged DNA is removed and repaired using opposite strand as template
Repair of double strand damage: a break in the DNA --> requires more error prone repair mechanisms.

question

What repair system is used when DNA damage is severe?

answer

a mechanism called the SOS regulatory system.
It's more error prone, which allows replication to proceed and the cell to replicate, but errors are most likely.

question

Translesion synthesis

answer

used by the SOS system, it allows DNA to be synthesized with no template.

question

RecA

answer

protein that recognizes and is activated by DNA damage detected as small bits of DNA that accumulate in the cell.
It interacts with the LexA repressor, which helps the LexA to cleave itself. It then falls off the opressor of the SOS regulons, which then can be transcribed.
It is also used during transformation by bringing strands of DNA closer together.

question

Radioduran

answer

an organism that is 20-200 times more resistant to radiation than E. coli. Why? it has hyper repair mechanisms.

question

Ames test

answer

Test makes practical use of bacterial mutations to detect for potentially mutagenic chemicals.

question

Genetic recombination

answer

Physical exchanged of DNA between genetic elements.

question

homologous recombination

answer

Process that results in the genetic exchange between two homologous DNA from two different sources. When two different alleles of the same gene are exchanged, one for the other.

question

Transformation

answer

genetic transfer process by which DNA is incorporated into a recipient cell and brings about genetic change.

question

What does it mean when a cell is competent?

answer

The the cell is capable of taking up DNA and being transformed.
electricity and chemical methods can be used to make cells more competent.

question

Transduction

answer

Transfer of DNA from one cell to another by bacteriophage. Basically, when a piece of the chromosome of the last victim of a virus is taken and given to the next "victim".
Two modes:
1) Generalized Transduction: DNA from any portion of the host genome is accidentally packaged inside the virion
2) Specialized Transduction: DNA from a specific region of the host chromosome is integrated directly into the virus genome.

question

Conjugation

answer

Mechanism of genetic transfer that involves cell-to-cell contact.
- plasmid encoded mechanism

question

What are essential features of conjugation?

answer

Donor cell and recipient cell.
F (fertility) plasmid - An episome that contains tra gene that encodes transfer functions.
Contains several transposable elements that allow the plasmid to integrate into the host chromosome. Encodes pilus.

question

What is the difference between Hfr, F+ cells.

answer

Cells possessing an integrated F plasmid are called Hfr (high frequency of recombination)
F+ - cells possessing non-integrated F plasmids

question

molecular cloning

answer

isolation and incorporation of a piece of DNA into a vector so it can be replicated and manipulated.

question

What are the main steps of gene cloning?

answer

1) Isolation and fragmentation of source DNA
- source DNA can be genomic DNA, RNA, or PCR-amplified fragments.
2) Insertion of DNA fragment into cloning vector
- vectors derived from plasmids or viruses. DNA ligase used to join two DNA molecules.
3) introduction of cloned DNA into host organism
- transformation is often used to get recombinant DNA into host.

question

Gene library

answer

Mixture of cells containing a variety of genes.

question

shotgun cloning

answer

gene libraries made by cloning random genome fragments

question

How do you detect the expression of a foreign gene with screening?

answer

antibodies
nucleic acid probes -- look for binding of labeled nucleic acid probe to DNA from specific colonies.

question

blue-white cloning -->

answer

the degradation of B-galactosidase creates a blue cell. you can screen for colonies that have the cloned vector in it by looking at the colors. Blue do not have the cloned vectors, white cells do.
the white cells are an example of insertional inactivation

question

Insertional inactivation.

answer

When a gene is inactivated by the insertion of foreign DNA.

question

Saccaromyces Cerevisiae

answer

Yeast cells

question

What are the molecular methods of mutagenesis?

answer

Site-directed mutagenesis
Casette mutagenesis and knockout mutations
- DNA fragment can be cut, excised and replaced by synthetic DNA fragment.
- Gene disruption is when the DNA is put into the middle of genes and causes a knockout mutation.

question

Expression vectors

answer

Allow experimenter to control the expression of a clone gene.
Based on transcriptional control and allows for high levels of protein expression.
Expression vectors are under the control of their own promotors and regulatory proteins.
IE in T7 expression vectors, cloned genes are placed under T7 promotors control. T7 RNA polymerase gene is present and recognizes only T7 promotors.

question

Virion

answer

A viral particle, the extracellular form of the virus.
Contains the N.A. genome surrounded by a protein coat.

question

What are the different viral classes and genomes?

answer

DNA viruses --> single stranded and double stranded DNA genomes.
RNA viruses --> single stranded and double stranded DNA genomes.
RNA <--> DNA viruses --> Single stranded RNA (Retroviruses) and Double Stranded DNA (hepadnaviruses)

question

How can viruses be classified?

answer

by the hosts that they infect.
Example: Bacterial viruses are bacteriophages,
archaeal viruses
animal viruses
plant viruses

question

What is the viral structure?

answer

Capsid --> the protein shell that surrounds the genome of a virus particle.
Capsomere --> subunit of the capsid. Continuously binds to RNA strand.
Nucleocapsid --> complete complex of nucleic acid and protein packaged in the virion
Enveloped virus --> virus that contains additional layers around the nucleocapsid.

question

What are the different symmetries of the nucleocapsids?

answer

helical symmetry: rod shaped viruses (tobacco mosaic virus)
Isohedral symmetry: Spherical viruses
Enveloped (budding) viruses: membrane surrounding the nucleocapsid.
Complex viruses: virions composed of several parts, each with separate shapes/symmetries. (isohedral heads and helical tails)

question

what is the structure of bacteriophage T4?

answer

From top to bottom:
Head, collar, tail, tail pins, end plate, tail fibers.

question

What enzymes do virions contain?

answer

lysozyme, used to bore a hole through the cell wall.
nucleic acid polymerases
neuraminidases - enzymes that cleave glycosidic bonds allowin the liberation of the virus from the host cell.

question

What are the different life cycles of viruses?

answer

virulent mode: viruses lyse host cells after infection
Temperate mode: viruses replicate their genomes in tandem with host genomes and without killing their host.

question

What type of lifecycle does bacteriophage lambda experience?

answer

temperate

question

What type of genome does bacteriophage T4 have?

answer

It has a dsDNA genome that is circularly permuted and terminally redundant.

question

T4 genome can be divided into three parts. What are they and what does each express?

answer

Early and middle proteins: Enzymes needed for DNA replication and transcription
Late protein: head and tail proteins and enzymes required to liberate mature phage particles.

question

Lysogen

answer

A bacterium containing a prophage

question

Lysogeny

answer

state where most virus genes are not expressed and virus genomes (prophage) is replicated in synchrony with host chromosome.

question

Lytic lifecycle

answer

Virus hijacks cells biosynthesis mechanisms until it bursts the cell with lysozyme

question

which two repressor proteins are key elements in the regulation of lytic vs. lysogenic events in lambda?

answer

cl protein (the lambda repressor) causes repression of lambda lytic events.
Cro repressor: controls activation of lytic events.

question

Animal viruses

answer

entire virion enters animal cell, unlike in prokaryotes.
Nucleus is site of replication for many viruses.
Much more likely to be enveloped.

question

what are the consequences of viral infections in animal cells?

answer

persistent infections: release of virions from host cell does not result in lysis
latent infections: delay between infection by the virus and lytic events.
transformation: conversion of normal cell into tumor cell
cell fusion: two or more cells become one cell with many nuclei

question

retroviruses

answer

viruses that replicate through a DNA intermediate.
- enveloped viruses
- contain reverse transcriptase (copies information from its RNA genome into DNA), integrase and protease.
Virion contains specific tRNA molecules.
Contains specific genes:
gag - encode structural proteins
pol - encode reverse transcriptase and integrate
env - encode envelope proteins

question

what is the process of replication for a retrovirus?

answer

-entrance into cell
-removal of virion envelope @ membrane
-reverse transcriptase of one or two RNA genomes
-integration of retroviral DNA into host genome
-transcription of retroviral DNA
-assembly and packaging of genomic RNA
-budding of enveloped virions; release from cell

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