Required practicals 3 and 4

Question 1

Calculate the molecular weight of KMnO4.
K = 39 g/mol
Mn = 55 g/mol
O = 16 g/mol

Answer 1

158g/mol

Question 2

Calculate the number of moles required to make 250 ml of 100 mM KMnO4.

Give your answer in moles, to two significant figures.

Answer 2

0.025 mol

Question 3

Now that you know how many moles of KMnO4 are required and you know the molecular mass of KMnO4 you should use your answers to the previous questions to calculate what mass of KMnO4 you require to make 250 ml of 100 mM solution.

Answer 3

3.95 g

Question 4

How much 100 mM stock solution is required to make 100ml of 100 micromolar solution?

Give answer to 2.s.f. in ml.

Answer 4

0.10 ml

Question 5

How do you dilute a concentrated acid?

Answer 5

Add the acid to the water.

Question 6

When diluting a concentrated acid, what happens to the solution?

Answer 6

It will get hot.

Question 7

When preparing a solution from NaOH pellets, what happens to the solution?

Answer 7

It will get hot.

Question 8

What volume of concentrated (11.64 M) HCl is required to make 100 ml of 1 M HCl?

Give answer to one decimal place in ml.

Answer 8

8.6 ml

Question 9

What should you do if you spill concentrated acid?

Answer 9

Rinse thoroughly with water.

Question 10

What was the electrophoresis gel made of?

Answer 10

Agarose.

Question 11

Why did we wear gloves to deal with KMnO4?

Answer 11

It is a powerful oxidising agent and will react quickly on skin. This will leave an insoluble brown stain of MnO2 which is not dangerous but will take a couple of days to fade.

Question 12

Why is the weighing boat rinsed into the beaker?

Answer 12

To ensure all KMnO4 is transferred.

Question 13

Why are the beaker and funnel also rinsed with distilled water?

Answer 13

To ensure complete transfer of MnO4.

Question 14

How was the stock solution/dilutions of KMnO4 solution disposed of?

Answer 14

Stock = waste bottle.
Dilutions = down the sink with the tap running (to make waste as inert as possible).

Question 15

Give the equation relating volume and concentration in terms of dilutions.

Answer 15

C1V1 = C2V2

(initial concentration x initial volume = new concentration x new volume)

Question 16

What must be done before making/modifying corrections to the electrophoresis tank?

Answer 16

The power supply is switched off.

Question 17

What must the buffer not spill over?

Answer 17

Leads and connections to the power supply.

Question 18

Name the components of a gel electrophoresis tank.

Answer 18

Sample wells
Anode
Cathode
Power supply
Buffer solution
Agarose gel

Question 19

Why is gel electrophoresis commonly used in molecular biology?

Answer 19

To separate DNA molecules, RNA molecules and protein molecules in complex mixtures according to their size and charge properties.

Question 20

How are the sizes of the separated molecules estimated?

Answer 20

When molecular weight markers are run alongside samples.

Question 21

When is agarose gel electrophoresis commonly used?

Answer 21

To separate DNA molecules.

Question 22

How are the DNA molecules separated?

Answer 22

Negative DNA molecules pulled through agarose matrix within an electric field.
Shorter molecules move faster than longer ones.

Question 23

List a property, other than the length of DNA molecules that is an important factor in migration rate.

Answer 23

The conformation (shape) of the DNA.

(Circular plasmids migrate the slowest and supercoiled move the fastest).

The voltage applied to the gel.

(Higher voltages give faster migration).

Question 24

How are small DNA molecules separated?

Answer 24

Increasing the concentration of agarose gel so the migration speed is reduced.

Question 25

Why must the voltage be limited?

Answer 25

It can heat up the gel and ultimately cause it to melt.

Question 26

How is DNA viewed?

Answer 26

DNA is stained with a marker which allows it to be visualised under UV light.

Question 27

Why must a centrifuge be balanced?

Answer 27

An unbalanced rotor can compromise the performance of your centrifuge, have an impact on the reliability of your testing results, and may even cause irreversible damage to your equipment.

Question 28

Why was the gel completely submerged with Tris-Borate-EBTA (TBE) buffer?

Answer 28

The buffer provides ions that carry a current through the gel, and to maintain a constant pH.

Question 29

How can you tell current is being produced?

Answer 29

Gas bubbles are visible from the cathode.

(Oxygen forms at anode and hydrogen at cathode).

Question 30

Why did we wear gloves for gel electrophoresis?

Answer 30

Gels are usually stained with a DNA stain, which can be toxic.

Question 31

Define cytotoxic (gel goes in cytotoxic waste bin).

Answer 31

Toxic to living cells.

Kills cells (including cancer cells).

Question 32

Why is it important to calibrate the electrode of the pH meter?

Answer 32

Regular Calibrations Ensure Accurate pH measurements.

Question 33

Comment on the accuracy of the acids and bases experiment.

Answer 33

Accurate:
- Regular calibrations of pH meter.
- Pellets of NaOH used instead of powder to
make 1 M and 0.1 M solution so all NaOH
transferred from weighing boat to
volumetric flask.
Not accurate:
- Only done once = results could be due to
chance.
- Smooth curve of best fit may not be the
best fit.
(Resulting in moles of HCl required to reach pH 7.40 being wrong and quoted pH range where HEPES solution acts as a buffer best).

Question 34

Why does the pH curve have an initial gradual decrease rather than a sharp one?

Answer 34

The HEPES solution is resisting pH change.

Question 35

Why is the curve flattest around pH 7-8?

Answer 35

HEPES molecules accept H+ from HCl.

Question 36

After pH 8, why does the curve gradually decrease again?

Answer 36

All HEPES molecules have bound to H+, so as more HCl is added there is excess H+ and pH decreases.