Quiz #4

Question 1

If a high concentration of NaCl were added to a hybridization solution, how would the stringency be affected?

Answer 1

Stringency would decrease

Question 2

Does an increase in temperature from 65C to 75C during hybridization raise or lower stringency?

Answer 2

Raise

Question 3

There are three methods to label a DNA probe. What is the name of the method, in which labeled nucleotides are added to the end of the probe using T4 polynucleotide kinase?

Answer 3

End labeling

Question 4

What is the definition of stringency?

Answer 4

Condition where two strands (RNA-RNA, DNA-RNA, DNA-DNA) hybridize

Question 5

To separate small proteins, would you use a 6? polyacrylamide gel or a 12% PA gel?

Answer 5

12

More PA means smaller pores

Question 6

For immunoblotting, to visualize the protein of interest, the membrane with the proteins is incubated first with a primary antibody against the protein of interest. Then, it is incubated with a secondary antibody against the primary antibody. That secondary antibody , for instance, has HRP conjugated to it. Why is HRP?

Answer 6

Horseradish peroxidase

Question 7

How does HRP work (in other words, what is one of the by-products that allow visualization of the protein of interest?

Answer 7

FINISH

Question 8

Activated cytotoxic T cells release two enzymes to disrupt the membranes of target cells and to tripper apoptosis. Name the two enzymes.

Answer 8

Granzyme - protease that chops up proteins

Perforin - creates a pore in the cell membrane to let granzyme into the cell

Question 9

What is the name of the dye used in Bradford assay in order to measure protein concentration?

Answer 9

Coomassie blue

Question 10

What wavelength of light is used to measure absorbance to measure protein concentration?

Answer 10

595 nm

Question 11

Tumor suppressor p53 (when there is DNA damage) will initiate two different pathways (one pathway involves p21) which are:

Answer 11

Apoptosis pathway (cell death) and non-apoptosis pathway (repairs damage

Question 12

Annexin V assay is used to detect what?

Answer 12

Early apoptotic cells

Question 13

How does the Annexing V assay work?

Answer 13

FINISH

Question 14

What is propidium iodide? What does it stain?

Answer 14

A stain/dye used to detect late apoptotic and necrotic cells. Ir can’t cross intact cell membranes. When the cell starts to rupture, it can pass through the openings in the membrane. It binds to DNA (DNA intercalator)

Question 15

A mutation whose gene product adversely affects the normal, wild-type gene product within the same cells, usually by dimerizing (combining) with it is known as a _______ mutation.

Answer 15

dominant-negative

Question 16

Describe “Sandwich ELISA”. You can draw a picture with description.

Answer 16

The wells of a plate are incubated with a primary “capture” antibody. The target protein is then added to the wells and incubated with another primary antibody. The target protein is then incubated with a secondary antibody (to the primary antibody). The secondary antibody is labeled with a radioactive or chemical marker. The marker allows visualization of the target protein.

Question 17

What is RT-PCR?

Answer 17

Reverse transcription polymerase chain reaction

Question 18

RT-PCR is used primarily for measuring _______.

Answer 18

mRNA

Question 19

FACS question

Answer 19

FINISH

Question 20

Modular nature of gene activator proteins (i.e. transcription factor) involves domains with specific functions. What is the name of one domain and its function?

Answer 20

Activation domain - recruits factors to activate transcription
DBD (DNA-binding domain) - binds DNA

Question 21

What is the difference between monoclonal and polyclonal antibody? What is the advantage of each type?

Answer 21

Monoclonal antibody - antibody derived from a single clone of a B cell; very specific; recognizes only one epitope; expensive; cleaner bands

Polyclonal antibody - antibody derived from multiple clones of a B cell; not as specific as monoclonal; recognizes all epitopes; strong signal, cheap; noisier bands