General Genetics Chapter 11 and 12

A primer bound to a template DNA strand is shown. What size fragments would be expected in the ddGTP tube?

10; 20; 22 nucleotides

After four rounds of replication there will be_______ copies of the target region and_______of these will only contain the double stranded target sequence.

16; 8

The Bam HI restriction site is often used in plasmid cloning. One strand of the site is shown below. What is the sequence of the sticky ends that will hybridize to a plasmid digested with Bam H1? 5'G↓GATCC

3'CTAG

The DNA sequence is read from the bottom of the gel toward the top. The resulting sequence will be:

5' to 3'

Two transgenic mice mate. What percentage of the progeny will have one copy of the transgene?

50%

What are the necessary components of a plasmid vector?

A gene encoding antibiotic resistance An ori A cloning site

What is the difference between a transcription factor and a coactivator?

A transcription factor directly binds to DNA and a coactivator does not.

Mutants that cannot convert _____ into cAMP cannot be induced to produce β-_____ because the concentration of cAMP is not great enough to activate the lac operon.

ATP; galactosidase

How do activator or repressor proteins function as genetic switches?

Activator or repressor proteins bind to operator sequences in the vicinity of the promoter to regulate transcription.

_____ or _____ proteins bind to operator sequences in the vicinity of the promoter to control its accessibility to RNA polymerase.

Activator; repressor

_____ cause operon structural genes to be expressed regardless of whether the inducer is present.

Constitutive mutations

GAL4 contains a(n) _________ domain that binds to the ________ on the _______ strand of GAL4-regulated genes.

DNA binding; UAS; coding

An operator mutation is a change in the

DNA sequence that is recognized in the lac repressor.

Four tubes are shown in the dideoxy sequencing reactions. What component is different for these four tubes?

Each tube has a different ddNTP (ddATP, ddCTP, ddGTP, or ddTTP).

Which of the following genes is constitutive?

GAL3

The clear mutants (designated by c) found by Francois Jacob in bacteriophage λ are analogous to the _____ and _____ mutants of the lac system.

I; O

The______ gene encodes the lac repressor protein which binds to the lac_________ in the ______ of lactose.

I; operator; absence

If the glucose in the environment was then depleted, how would the lac operon mRNA production change?

Increased production

An operon is a group of genes under the control of a single promoter. Match each type of operon with the descriptions. Inducible Repressible Constitutive

Inducible: Under default conditions, transcription is inhibited Repressible: Transcription stops when the repressor gene product is activated. Sufficient amounts of the gene product inhibit further transcription. Constitutive: Transcription occurs regardless of environmental conditions.

How much lac operon mRNA does the lac operon produce when both lactose and glucose are present?

Low mRNA production

Tryptophan is an amino acid necessary for E. coli survival and growth. E. coli contain genes coding for enzymes that synthesize tryptophan. These genes are grouped together on a segment of DNA called the tryptophan (trp) operon. Cells can use these enzymes to synthesize tryptophan when it is not present in the environment. However, when tryptophan is already present in the environment, cellular resources are shifted away from manufacturing the enzymes for tryptophan synthesis. Tryptophan binds with and activates the trp repressor, which then binds to the trp promoter and blocks RNA polymerase. Blocking RNA polymerase decreases the normal transcription rate of the operon. What type of regulation does the trp operon exhibit?

Negative regulation

Which of these are constitutive mutants of the lac system and as such would cause the constant expression of the Z, Y, and A genes associated with the lac operon?

O^C and I−

What is the consequence of parental imprinting?

Only one copy of the gene is expressed.

How is dosage compensation demonstrated in Drosophila?

The X‑chromosome is transcriptionally upregulated two‑fold in males.

A larger protein will bind more of the anionic SDS detergent than a smaller protein, yet it migrates more slowly through the polyacrylamide network toward the + electrode. Why doesn't the protein with more negative charge move faster toward the + electrode?

The charge/mass ratio is constant, and the proteins migrate according to mass.

What occurs as a result of allolactose binding to a repressor protein?

The repressor falls off the DNA, allowing RNA polymerase to begin transcription.

Why does Taq polymerase need to be stable at high temperatures for successful PCR?

The sample must be heated to 95 degrees Celcius to break H-bonds holding the DNA strands together so that primers can anneal at each cycle.

During transformation, only one plasmid enters an E. coli cell that is competent to take up DNA. What could happen if multiple plasmids entered one E. coli cell?

There could be a mixture of different plasmids in each colony.

A bacterial colony is grown in media containing a low level of glucose. Which of these statements is TRUE? There is a low level of cAMP in these cells. There is a high level of cAMP in these cells. The amount of cAMP in these cells is equal to that of cells grown in high-glucose conditions. There is no cAMP in these cells.

There is a high level of cAMP in these cells.

Which is not a key regulatory component of the lactose metabolic system? binding site for regulatory protein binding site for RNA polymerase transcription regulatory gene a constitutive mutation

a constitutive mutation

What is an operator?

a short sequence of DNA located near the promoter region that is recognized by a repressor protein

What is an inducer?

a small molecule that activates transcription by inhibiting the action of a repressor protein

In the cytoplasm, some of the lactose is converted into _______ , which binds the _______ site of the lac repressor.

allolactose; allosteric site

Regulatory proteins typically contain sites that can bind to other proteins or small molecules. Through such binding interactions, these sites can cause a repressor or activator to be inactivated or become functional. These are _____ sites.

allosteric

A(n)_____ is a slight alteration in the shape of a repressor protein that alters the DNA-binding site so that the repressor no longer has high affinity for the operator.

allosteric transition

What does constitutive mean in regards to the lac operon?

always on

What is an insulator?

an element that disrupts the signal between enhancers and promoters

A ddGTP is a chain terminator because it is incorporated opposite a ________ in the growing DNA chain, but lacks a ________.

cytosine; 3'-OH

Which of the following is NOT a function of SDS during SDS-PAGE? denatures RNA disassociates subunits of multi-subunit proteins binds uniformly to polypeptides resulting in a constant charge/ mass ratio unfolds proteins allows all proteins to move toward the + electrical charge

denatures RNA

The two conformations of the AraC protein are dependent on whether the allosteric _____, arabinose, has bound to the protein.

effector

Which chromatin structure allows genes to be transcriptionally active?

euchromatin

IS mutantations turn _______ the lac operon in the presence of lactose and turn ________the lac operon in the absence of lactose.

off; off

The lac operon in an I-O+Z+Y+ allele will be _______in the presence of lactose and _______ in the absence of lactose.

on; on

A researcher wants to make a mouse with a gene harboring a dominant mutation (polydactyly) that causes extra fingers and toes to develop in humans. A clone of the human gene is obtained and injected into:

one pronucleus of a fertilized egg.

What is associated with unmethylated CpG islands at promoters?

open chromatin and active transcription

Which group of proteins work in opposition to maintain the repressed and active transcription states of parent cells in daughter cells?

polycomb and trithorax

What is a result of chromosomal rearrangement?

position‑effect variegation

What defines the biological specificity of gene regulation?

the chemical interaction between regulatory proteins and DNA sequences

Below is a list of steps for performing a Northern blot. Place them in the correct order. 1. Size fractionate RNA by gel electrophoresis 2. Expose membrane to X-ray film 3. Wash off probe that is not bound by complementary base pairing. 4. Isolate cellular RNA 5. Incubate membrane with radioactive GAL gene 6. Grow yeast in galactose or glucose medium 7. Transfer RNA to an absorbent membrane by capillary action

6 4 1 7 5 3 2

If "X" is cAMP, what is "Y"?

CAP

What enzyme is introduced into a cell along with a singe guide RNA?

Cas9

Why is the design of single guide RNA limited to sequences that are next to PAMs?

Cas9 recognizes PAM sites to create double strand breaks.

_____ inhibit(s) the synthesis of digestive enzymes.

Catabolite repression

Which of the statements describe how inducers cause changes in gene expression?

Inducers bind to repressor proteins and prevent the repressor from inhibiting transcription. Inducers allow transcription of specific genes.

Which of the following genes would be expected to work in trans (on a different DNA molecule)?

a gene encoding a DNA-binding protein

What is a regulatory gene?

a gene that controls the expression of one or more genes by promoting or inhibiting transcription

What is a repressor?

a regulatory protein that blocks transcription by binding to the operator site

Walter Gilbert and Benno Müller-Hill demonstrated all of these EXCEPT: a repressor-bound region of DNA in the lac operon (i.e., the operator) is damaged by DNase treatment. the operator of the lac operon is actually transcribed and represented in the mRNA transcript. point mutations in the operator of the lac operon can interfere with repressor-operator interactions. the Lac repressor contains four distinct sites to which lactose can bind.

a repressor-bound region of DNA in the lac operon (i.e., the operator) is damaged by DNase treatment.

Bacteria are nutritional opportunists and, as such, they are able to do all of these things EXCEPT: recognize nutrients in the environment. repress the expression of relevant genes when needed. break down any substance into digestible pieces activate the expression of relevant genes when needed.

break down any substance into digestible pieces

If resources are limited, _____ is more active and more _____ is produced.

cII; λ repressor

The GAL gene is transcribed into mRNA when yeast cells are grown on galactose. What type of role might the GAL gene have?

catabolic or galactose transport

Looking at the genetic map, we see that from PR, _____ is the first gene transcribed, and from PL, _____ is the first gene transcribed.

cro; N

Which environmental signal facilitates the regulation of GAL4?

galactose

Which is not located within a bacterial operon? operator regions that bind to repressors genes for different metabolic pathways promoter regions that bind to RNA polymerase genes for the same metabolic pathway

genes for different metabolic pathways

Catabolite repression refers to the repression of the lac operon in the presence of _____.

glucose

It is more efficient for the cell to metabolize _____ rather than _____.

glucose; lactose

A researcher is growing E. coli in nutrient-rich media. He infects these bacteria with the λ phage and treats these cells with ultraviolet light. In these bacteria, the λ phage most likely:

has entered the lytic cycle.

The injected fertilized eggs are______ for the transgene and injected into a foster mother. Ten to thirty percent of progeny are _______ for the transgene in every tissue.

heterozygous; heterozygous

What repair mechanism allows for the generation of precise mutations?

homologous recombination

A graduate student is studying the expression of the AraC protein in bacteria grown in arabinose-poor conditions. She notices that in these cells, AraC always associates simultaneously with two distinct regions on the ara operon. These regions are the _____ and the _____.

initiator; operator

IS mutant alleles encode lac repressors that are unable to bind to

lactose

Where will an introduced gene integrate into the mouse genome?

nearly anywhere since integration is random

Bacteria are different from eukaryotes in all of these ways EXCEPT in their _____. need to regulate gene expression metabolism cellular structure size

need to regulate gene expression

After SDS-PAGE, the gel is placed next to nitrocellulose. Where will the + charge need to be to get the proteins to migrate to the nitrocellulose? Why is nitrocellulose used?

next to the nitrocellulose; nitrocellulose binds proteins

The N gene encodes for the _____ and promotes _____.

positive regulator; cII and cIII expression

ISO+Z+Y+/F'I+ alleles will result in________ of the lac operon in the presence of lactose and _______of the lac operon in the absence of lactose, because the IS mutation is ________ to I+.

repression; repression; dominant

Under stress, the bacterium forms _____ that are remarkably resistant to heat and desiccation.

spores

RNA polymerase cannot transcribe the lac operon when?

the lac repressor is bound to the operator.

What is operon induction?

the relief of repression systems

Which observation demonstrates attenuation as a bacterial control mechanism? the breakage of lactose into glucose and galactose the removal of tryptophan decreases mRNA expression the simultaneous induction of coordinately controlled genes the removal of tryptophan increases mRNA expression

the removal of tryptophan increases mRNA expression

What is the role of chromatin in transcriptional regulation?

to act as a substrate for reversible changes

An F' plasmid with an I+ gene is introduced into the I-O+Z+Y+ allele. The I+ gene operates in _______ and is _______ to I-.

trans; dominant

A researcher wants to make a mouse with a gene harboring a dominant mutation (polydactyly) that causes extra fingers and toes to develop in humans. Transgenic mice with the introduced genes are produced. These mice appear normal and have normal numbers of toes on their forelimbs and hindlimbs. What should the researcher test before deciding the gene affects humans differently than mice?

Expression levels of the transgenic mRNA and protein.

Classify the given examples of prokaryotic gene expression as positive or negative gene regulation.

Positive Gene Regulation: In the presence of lactose and low glucose, the lac operon is expressed 20-fold higher than in the absence of lactose. In the presence of the sugar arabinose, an activator protein binds the promoter of the genes responsible for processing arabinose and induces their transcription. Negative Gene Regulation: In the presence of iron, the dtxR repressor protein binds DNA, and the gene that encodes for the diphtheria toxin is not expressed. In the absence of lactose, the lacR repressor protein binds the lac operon. In the presence of excess tryptophan, a repressor protein binds the operator of the trp operon and prevents the operon from being transcribed.

In the presence of arabinose, both the CAP-cAMP complex and the AraC-arabinose complex must bind to _____ in order for RNA polymerase to bind to the promoter and transcribe _____.

araI; araO

An Oc mutation results in transcription of the lac operon in the _______ of lactose.

presence and absence

Which DNA segment binds to RNA polymerase for transcription? promoter operon enhancer operator

promoter

Genetic switches are _____ and _____ that control gene transcription.

proteins; DNA sequences

The IS mutation studied by Jacob and Monod demonstrated several interesting characteristics: it was dominant, resulted in the constant repression of the lac operon, and was able to repress both lac operons in partial diploid bacteria. Thus, IS could be considered a _____ mutation.

superrepressor and trans-acting

Which of the following is not a component of a PCR reaction? template genomic DNA nucleotides template RNA Taq polymerase DNA primers

template RNA

The lac operon contains three genes: lacZ, lacY, and lacA for β‑galactosidase, galactoside permease, and thiogalactoside transacetylase, respectively. The operon also contains a promoter site and an operator site. The regulatory gene includes the I gene for the Lac repressor protein and a promoter for that gene. The lac operon contains three genes: lacZ, lacY, and lacA for β‑galactosidase, galactoside permease, and thiogalactoside transacetylase, respectively. The operon also contains a promoter site and an operator site. The regulatory gene includes the I gene for the Lac repressor protein and a promoter for that gene. Determine which events will increase the concentration of lac gene products and which will decrease the concentration of lac gene products.

Increase in Gene Products: The I gene is disabled Lactose is present in the cell Allolactose binds to the Lac repressor protein The Lac repressor dissociates from DNA Decrease in Gene Products: Allolactose is not present in the cell The Lac repressor remains bound to the operator

What is a promoter?

a sequence of DNA located upstream of a gene that signals the start of transcription

Which of these statements is TRUE? An OC mutation in the lac operon can affect the expression of structural genes on an F′ factor. An OC mutation can prevent the expression of the diffusible Lac repressor. A haploid bacterium with the genotype OCZ+Y+ would never express permease or β-galactosidase. An OC mutation on the lac operon is a cis-acting mutation.

An OC mutation on the lac operon is a cis-acting mutation.

Which is an example of negative regulation? Low glucose levels promote formation of the CAP‐cAMP complex. The CAP‐cAMP system acts as a signal to induce expression. Large amounts of mRNA are produced only when lactose is present. An active repressor binds to an operator site.

An active repressor binds to an operator site.

Classify each of the characteristics as pertaining to gene regulation in either prokaryotes or eukaryotes.

Prokaryotic Gene Regulation: DNA does not contain introns Some genes are organized into operons, and mRNA transcripts often specify more than one protein mRNA can be transcribed from DNA and translated into protein at the same time Eukaryotic Gene Regulation: Genes are located on different chromosomes A 5' cap and a 3' poly-A tail are added to the mRNA Transcription occurs in the nucleus, whereas translation occurs in the cytoplasm

Which one of these statements is INCORRECT? Glucose, the preferred energy source for E. coli, represses genes involved in the metabolism of other sugars. The CAP protein is involved in catabolite repression of the lac operon and is the portion of the CAP-cAMP complex that actually binds to DNA. As part of catabolite repression, high glucose concentrations decrease the amount of cAMP in bacterial cells. RNA polymerase is the effector molecule that enables CAP to bind to the lac operon promoter.

RNA polymerase is the effector molecule that enables CAP to bind to the lac operon promoter.

An I- mutation is unable to?

bind the operator DNA.

A researcher wants to create a mutant strain of bacteria that fails to transcribe the ara operon, even in the presence of arabinose and the absence of glucose. To achieve this end, the researcher could induce mutations in all of these EXCEPT: the O site of the ara operon. the I site of the ara operon. the gene encoding AraC. the gene encoding CAP.

the O site of the ara operon.

A researcher has identified a possible repressor in a new species of bacteria. She notices that for its associated gene, the repressor always binds to the same DNA region just downstream of the activator and promoter regions. The region of DNA to which this repressor binds is the _____.

operator