Molecular Biology Answers

Question 10: Why do you think the protocol does not say to dissolve compounds directly in 1 liter of water?

Question 11: The PBS protocol above says to adjust pH to 7.4 with HCl. What does this imply on the pH of 10x PBS before adjusting the pH, would it be greater or smaller than 7.4?

Question 12: The last step in the protocol is to autoclave the 10x PBS solution. Why do you think this step is important? Look up the definition of autoclave if you are unsure what it means.

Question 13: Taking into account your response to question 5, now that you have made a 10x PBS solution, describe how you would prepare 1 liter of 1x working solution PBS, including which glassware you would use. Will you need to adjust the pH again?

Question 6: What is the first thing to do after putting a weighing boat on the balance?

Question 7: If you have excess reagent on the weighing boat, what should you avoid doing and why?

Question 8: If you had the choice between a 1 liter beaker and a 1 liter graduated cylinder, which one should you use to measure volumes with maximal precision when making 1 liter of PBS? (you can perform an internet search to find this if you are not sure of the answer)

Question 9: What should be done before measuring an unknown pH of a solution using a pH meter?

Question 5: As previously stated, the concentration of NaCl, KCl and Phosphate in working strength 1 x PBS is 0.137M NaCl, 0.012M Phosphate, 0.0027M KCl, pH 7.4 How do they compare to the concentrations you calculated for 10x PBS?

Question 4: Using periodic table found in your textbook, calculate (to 2 decimal places) the molecular mass for each of the compounds used to make PBS.

Create the following table and fill it in with the mass of each component required to make 1 liter of 10 x PBS (the recipe for 10x PBS is below question 2) and their final molar concentration in the buffer calculated as described above.

Compound formula Molecular mass (in g/mol) Mass of compound per liter of 10x PBS (in g) Molar concentration (in mol/l)
NaCl
KCl
Na2HPO4
KH2PO4

Question 2: Based on the equation below, which ion plays the role of hydrogen-ion donor (acid) and which ion plays the role of hydrogen-ion acceptor (base) in PBS?
H2PO4-(aq) ⇆ H+(aq) + HPO42-(

The composition of PBS is 0.137M NaCl, 0.012M Phosphate, 0.0027M KCl, pH 7.4. Below is the protocol to make 1 liter of 10x concentrate PBS.
Combine the following:
• 80g NaCl
• 2g KCl
• 14.4g Na2HPO4 (dibasic anhydrous)
• 2.4g KH2PO4 (monobasic anhydrous)
• 800mL distilled H2O

1. Adjust pH to 7.4 with HCl
2. Add H2O to 1L
3. Autoclave for 20 minutes on liquid cycle. Store at room temperature.

Question 13: Taking into account your response to question 5, now that you have made a 10x PBS solution, describe how you would prepare 1 liter of 1x working solution PBS, including which glassware you would use. Will you need to adjust the pH again?

Question 12: The last step in the protocol is to autoclave the 10x PBS solution. Why do you think this step is important? Look up the definition of autoclave if you are unsure what it means.

Question 11: The PBS protocol above says to adjust pH to 7.4 with HCl. What does this imply on the pH of 10x PBS before adjusting the pH, would it be greater or smaller than 7.4?

The recipe for PBS says to dissolve compounds in 800 ml of water, adjust the pH to 7.4, then add water up to 1 liter. The final pH should still be 7.4, because the pH of buffer solutions remains stable when they are diluted as long as the concentration of its constitutive acid and base is not too low.

Question 10: Why do you think the protocol does not say to dissolve compounds directly in 1 liter of water?

Question 9: What should be done before measuring an unknown pH of a solution using a pH meter?