Techniques in experimental biology (do not need to know).

Question 1

What is the definition of electrophoresis?

Answer 1

’ The movement of charged particles in fluid or gel under the influence of an electric field.’

Question 2

When was the technique of electrophoresis established?

Answer 2

1970’s.

Question 3

In electrophoresis are small or large molecules separated largely independent of size?

Answer 3

Small.

Question 4

What charge do RNA molecules generally have?

Answer 4

Negative with positive patches.

Question 5

What molecule is bound to all RNA’s?

Answer 5

Mg2+.

Question 6

Some RNA molecules can act as catalysts. What does this mean they can be classed as?

Answer 6

Metalloenzymes.

Question 7

Why is the shape of RNA more complex than DNA?

Answer 7

RNA is bent.

Question 8

What end of the gel do all proteins and DNA go to in electrophoresis?

Answer 8

Anode.

Question 9

What is lost in electrophoresis when the voltage is turned of?

Answer 9

Resolution/ clarity. This is because the proteins/DNA diffuse after roughly five minutes.

Question 10

How many volts are used in electrophoresis?

Answer 10

100.

Question 11

Does protein or DNA electorphoresis cost more?

Answer 11

Protein.

Question 12

What two key words can describe electrophoresis?

Answer 12

POWERFUL and CHEAP.

Question 13

In electrophoresis the loading buffer is prepared as what?

Answer 13

Concentrates of the concerntrate sample, for example 2 or 5x.

Question 14

What is the purpose of the buffer component in the loading buffer in electrophoresis?

Answer 14

It supports and stabilizes ions.

Question 15

What charge does the inert dye in a loading buffer have and why?

Answer 15

Negative as it is the same charge as the molecule.

Question 16

Loading buffers contain a ________ component.

Answer 16

Viscous.

Question 17

The viscous component normally used in loading buffers is glycerol. Sucrose can also be used. What is the downside of using sucrose?

Answer 17

It can be degraded by bacteria and therefore go of.

Question 18

A denaturant is often added to RNA and proteins in electrophoresis. Which is it more important in?

Answer 18

Proteins.

Question 19

What denaturant is used in protein electrophoresis?

Answer 19

SDS.

Question 20

The loading buffer used with proteins in electrophoresis has a neutral pH and low ionic strength. What is it called?

Answer 20

Tris.

Question 21

Is the pH of the loading buffer higher in protein or DNA electrophoresis?

Answer 21

DNA.

Question 22

What are Bromophenol blue and Xylene cyanol?

Answer 22

Dyes used in the loading buffer for DNA electrophoresis.

Question 23

What type of electrophoresis is the loading buffer Tris- acetate- EDTA used in?

Answer 23

DNA.

Question 24

What chemical is used as the denaturant in RNA electrophoresis?

Answer 24

Formanide.

Question 25

SDS is _____ charged. The amino acid chain ____ out as the ______ component sticks to the polypeptide _______. This results in all polypeptide chains being ______ ______ negatively charged rods.

Answer 25

Negatively, straightens, hydrophobic, different sized.

Question 26

What is B-Mercaptoethanol used for, especially with antibodies?

Answer 26

To oxidize disulphide bridges so they do not bunch up meaning the electrophoresis gel can still run.

Question 27

2000 proteins can be found in one eukaroyotic cell. How many are normally found at one one?

Answer 27

5000-6000.

Question 28

Although there are 2000 possible proteins found in one eukaryotic cell how comes 60,000 can theoretically be found?

Answer 28

Post transcriptional modifications.

Question 29

How many proteins can you separate on an SDS phage gel?

Answer 29

200.

Question 30

What gel is found on top in a SDS gel?

Answer 30

The stacking gel. The resolving gel is found below.

Question 31

What polymer is found in the stacking gel which forms cross linkages when added to the bicacrylamide cross linker?

Answer 31

Acrylamide long chain.

Question 32

What is the purpose of the stacking gel in the SDS gel?

Answer 32

Aligns proteins.

Question 33

Can you run SDS gels without the stacking or the resolving gel?

Answer 33

Stacking.

Question 34

What percentage of crosslinking in the SDS gel will prevent proteins from moving?

Answer 34

20%/

Question 35

Is acrylamide or bicacrylamide a neurotoxin?

Answer 35

Acrylamide.

Question 36

Acrylamide is innert when it is polymerised. This process takes 5 minutes. What two other things are needed in thus process?

Answer 36

APS and TEMED amine.

Question 37

What is the pH of the stacking gel?

Answer 37

6.8.

Question 38

What is the pH of the resolving gel?

Answer 38

8.8.

Question 39

What process is good at identifying different proteins in mixtures?

Answer 39

Western blotting.

Question 40

What material is the filter usually made from in Western blotting?

Answer 40

Nylon.

Question 41

What orientation are the agarose gels when nucleic acids are being run?

Answer 41

Horizontal.

Question 42

Nucleic acid electrophoresis is more simple then protein electrophoresis. What two companents are needed?

Answer 42

Agarose and buffer which are then boiled together.

Question 43

Was nucleic acid or protein electrophoresis discovered first?

Answer 43

Protein. Nucleic acid electrophoresis was discovered 10 years later.

Question 44

What type of blot is used with RNA?

Answer 44

Northern.

Question 45

What type of blot is used with DNA?

Answer 45

Southern.

Question 46

By separating by charge, size and by using two orientations more proteins can be separated with an SDS gel, although this is not usually used anymore. How many proteins can be separated?

Answer 46

1000 instead of 200.