Final Lab Review: Cell Tissues, Gene Expression, and Cloning Techniques

Which of the following tissues originates from the mesoderm?

Muscle

The retina contains which of the following cell types?

Photoreceptors

The eye develops from the ______.

ectoderm

True or False: All genes are expressed in all cells at all times.

False

What does a darker color indicate in a heatmap of gene expression?

Higher expression

General transcription factors are required by ______ genes.

all

Pax6a is considered a master regulator because:

It can induce eye formation in ectopic locations

True or False: Pax6a and Pax2a are co-expressed in the same retinal cells.

False

What happens when Pax6a is lost in eye development?

Failure of normal eye formation, reduced or absent

RNA samples are kept at ______ °C to prevent degradation.

-80

Why did we clean with hydrogen peroxide?

To degrade RNases

Why should face masks be used when working with RNA?

To prevent contamination by RNases from breath

What chemical in Trizol can cause burns?

Phenol

During organic extraction, RNA is found in the ______ phase due to its polarity.

aqueous

True or False: Chloroform was used to dissolve the RNA pellet.

False

True or False: RNA is more stable than DNA.

False

Which enzyme degrades RNA?

RNase

To reduce RNA degradation, we used _________ on surfaces and tools.

RNAase decontaminating solution

What is the main function of Trizol?

Protein and RNA separator

True or False: Trizol contains phenol and is toxic to skin.

True

The _______ layer contains denatured proteins.

organic

What is a chromatogram, and what is its function in DNA sequencing?

A graphical output that displays the results of DNA sequencing by showing colored peaks for each nucleotide

True or False: Reverse transcriptase converts cDNA into RNA.

False

Which primer is used to bind to the mRNA tail?

Oligo dT

Which is NOT a component of the RT reaction?

Taq polymerase

True or False: Ice is used to stabilize RNA before cDNA synthesis.

True

RT reactions are incubated first at ______ °C to anneal primers.

25

What is the role of primers in PCR?

Define the region to be amplified

True or False: Primers are RNA sequences.

False

Forward and reverse primers bind to ________ DNA strands.

complementary

What temperature is used for primer annealing in PCR?

54°C

True or False: PCR consists of 3 temperature steps: denature, anneal, extend.

True

The extension phase typically occurs at ______ °C.

74

What is special about Taq polymerase?

It is heat-stable

True or False: Taq polymerase adds adenine overhangs for TA cloning.

True

Taq comes from the bacterium _______ aquaticus.

thermus

What does a band on the gel represent?

Amplified cDNA

True or False: You always know the exact identity of the PCR band without sequencing.

False

A DNA ladder is used to estimate the band's ______.

size

Why do we use Pax6a as a control?

It confirms PCR reagents work

True or False: A positive control helps validate negative results.

True

Pax6a is a known ________ regulator of eye development.

master

True or False: TA cloning requires restriction enzymes.

False

A vector for TA cloning has a 3' ______ overhang.

T

What does the AmpR gene do?

Confers antibiotic resistance

True or False: Only transformed bacteria survive on Amp plates.

True

______ cells are plated on Amp plates after transformation.

transformed bacterial

What causes a colony to appear white?

Inserted DNA disrupts lacZ

True or False: Blue colonies lack the insert.

True

X-gal turns blue when cleaved by ________.

B galactosidase

What is the role of IPTG in blue/white selection?

Induces lacZ expression

True or False: IPTG is a digestible lactose analog.

False

IPTG activates transcription of the ______ operon.

lac

What is isolated during a miniprep?

Plasmid DNA

True or False: Miniprep uses spin column purification.

True

The resuspension solution helps _______ the cells.

resuspend

What does the lysis buffer do?

Breaks open cells

True or False: Vortexing during lysis can shear chromosomal DNA.

True

Neutralization causes chromosomal DNA to form a ______.

precipitate

What does EcoRI do?

Cuts at specific sites

True or False: Two bands after EcoRI digest suggest successful insertion.

True

EcoRI is a type ______ restriction enzyme.

II

What does a gel confirm after digestion?

Insert presence and size

True or False: A single band may suggest vector re-circularization.

True

DNA migrates toward the ______ electrode in a gel.

positive

What is a multiple cloning site?

A region with many restriction sites

True or False: Insertion within MCS disrupts blue colony formation.

True

A plasmid map helps locate _______ for cloning.

restriction sites

Why do we sequence the insert?

To confirm insert identity

True or False: Sanger sequencing uses chain-terminating nucleotides.

True

Successfully cloned inserts will have matching _______ with the target gene.

sequence

Still learning (3)

You've begun learning these terms. Keep up the good work!