EXAM: What I still don't know

Question 1

What enzyme relieves supercoiling?

Answer 1

DNA topoisomerases

Question 2

What creates 5’ cap?

Answer 2

Capping proteins

Question 3

What proteins bind to make poly A tail and cleave?

Answer 3

Cleavage and polyadenylation factors (CPSF, CstF)

Question 4

Error rate of tRNA synthetase?

Answer 4

1/40,000

Question 5

First thing initiator tRNA/small subunit complex does after binding?

Answer 5

Finds AUG site

Question 6

What stops translation at the very end?

Answer 6

Release factors that bind and dissociate everything

Question 7

What repressor adds with -lactose in LAC operon?

Answer 7

Lac repressor

Question 8

What activator adds with -glucose in LAC operon?

Answer 8

CAP with cyclic AMP to activate

Question 9

What transcription factor binds to TATA box in transcription initiation?

Answer 9

TBP with TFIID

Question 10

What causes patterns of histone modification as the result of activator proteins?

Answer 10

histone modifying enzyme

Question 11

What protein might cause mRNA to decay before translation and how?

Answer 11

deadenylase, binds to 5’ cap and eats tail

Question 12

What protein might cause mRNA to decay before translation and how?

Answer 12

deadenylase, binds to 5’ cap and eats tail

Question 13

What protein and system is involved in gene silencing via heterochromatin formation?

Answer 13

siRNAs binding to RITS, cause methylation and stops genes from going

Question 14

What protein and system is involved in gene silencing via heterochromatin formation?

Answer 14

siRNAs binding to RITS, cause methylation and stops genes from going

Question 15

I’m in the lab and I want to only grab C. el neuronal cells. What do I do?

Answer 15

Use FACS (Fluorescence-activated cell sorter). For separating cells from other tissues.

Question 16

How would FACS work to grab C. elegans neurons?

Answer 16

• Bind antibody with a fluorescent dye to surface of neurons. Dye makes them neg charged
• They go down a tube. A laser shoots it
• Further down the tube, there are two electrical fields at opposing charges
• These separate now-negative neurons from positive other cells

Question 17

Now I want to take those neurons and extract their nuclei. How do I start that?

Answer 17

Put test tube in a centrifuge, make it go

Question 18

Now I want to take those neurons and extract NMDAR. How do I start that?

Answer 18

Put test tube in a centrifuge, make it go fast

Question 19

I have C elegans and got a test tube with only proteins in it from centrifugation. I have a bunch of NMDAR already. How do I separate them?

Answer 19

Column chromatography

Question 20

What do I tag the C. elegans proteins with to separate them with column chromatography?

Answer 20

Peptide epitope tags

Question 21

What two chemicals do I have in front of my SDS page? Why?

Answer 21

SDS - to make my NMDARs negatively charged so they go down gel

B-mercaptoethanol - to break disulfide bonds in NMDARs

Question 22

In my SDS page, how could proteins be negatively charged with SDS and still move along a charge gradient?

Answer 22

Makes some proteins more negatively charged than others?

Question 23

What would I stain agarose gel with to see the NMDAR gene? What kinds of gels could I use?

Answer 23

Agarose gel to lots (1000) of nucleotide pairs

Polyacrylamide gel to see 50 nucleotides of the gene

Pulsed field gel to see entire chromosomes

Question 24

How would I make the NMDAR gene radioactive to see where it is?

Answer 24

In situ hybridization: add 32p-labeled nucleotides, make polymerase duplicate using them

Question 25

I’m making cDNA out of NMDAR. What do I need?

Answer 25

Poly-T polymerase for template

RNAse to digest RNA after first cDNA strand

DNA polymerase to make cDNA strand

Question 26

What’s the electron microscope with worse pictures? What’s the advantage?

Answer 26

TEM (transmission electron microscope)

Can go through things to see the inside

Question 27

What are the large channels comprising porins made from?

Answer 27

beta barrels form them

Question 28

How do you grab an individual section of bilayer?

Answer 28

Use detergents to disrupt and bring out

Question 29

What are lipid-detergent groups called?

Answer 29

micelles

Question 30

How would you remove an NMDAR receptor and put it into another membrane?

Answer 30

Use detergent that surrounds it then takes it out