PAG 7 Microbiological Techniques: OCR A Level Biology Exam Questions by Topic

OCRPAGPractical SkillsExam Questions
Written by Tom Whitburn

OCR A PAG 7 · 15 questions · mark schemes and examiners’ tips included

PAG 7 is about growing microorganisms safely and counting them. You will have used aseptic technique to inoculate nutrient broth or agar plates, incubated cultures at a safe temperature, and estimated the size of a bacterial population by serial dilution, plating and counting colonies.

The examiners see the same slips every year. Students write 'amount' instead of naming a variable, say 'control' without explaining what it checks, or think the Bunsen flame kills every microbe. In the calculations, marks go on tracking each dilution factor, scaling up from the volume plated to the whole culture, and giving the answer in standard form. Choose a plate with plenty of separate, countable colonies.

Work through the fifteen questions below in order. They start with aseptic technique, move on to controlling a growth investigation, and finish with serial dilutions and colony count calculations. If you'd like to go through questions like these with me each week, my OCR A masterclasses run on Wednesdays at 5.15pm for Year 12 and on Tuesdays at 6.30pm or Thursdays at 5.15pm for Year 13.

What these questions cover (OCR PAG 7: microbiological techniques)

PAG 7 is one of the twelve practical activity groups in the OCR A Level Biology A specification (H420). The questions test these parts of the specification:

  • 1.1.1(b) identification of variables that must be controlled, where appropriate
  • 1.1.4(e) the refining of experimental design by suggestion of improvements to the procedures and apparatus
  • 1.2.2(c) use of laboratory glassware apparatus for a variety of experimental techniques to include serial dilutions
  • 1.2.2(i) use of microbiological aseptic techniques, including the use of agar plates and broth
  • 6.2.1(g)(i) how to culture microorganisms effectively, using aseptic techniques
  • 6.2.1(h)(ii) practical investigations into the factors affecting the growth of microorganisms (including estimating population size from colony counts)

How to use this page

Write your answer on paper before you open the mark scheme. Allow about 1.2 minutes per mark: the 15 questions are worth 29 marks, so about 35 minutes in total. Then mark your answer strictly against the mark scheme and read the examiners' tip underneath it. If you'd rather print the questions, download the PDF pack.

Aseptic technique

Question 1

A level Paper 2 · June 2022 · Q17(a)(i) · 1 mark · Easy

State why the technique for culturing microorganisms must be aseptic.

Question 17: students divide a large bacterial culture into smaller 50 cm³ flasks, incubate them at 20 °C for up to 48 h and count total and viable bacteria every 4 h; part (a)(i) asks why culturing must be aseptic.
Show the mark scheme Mark scheme: to prevent contamination by unwanted microorganisms, or prevent their entry or growth; 'kill' is ignored.

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From the examiners' report: Nearly everyone got this. Use the word contamination, or say it stops unwanted microorganisms getting in or growing.

Question 2

A level Paper 3 · June 2017 · Q2(b)(ii) · 2 marks · Medium

Describe two practical considerations for establishing a yeast culture successfully.

Part (b)(ii) asking for two practical considerations to make sure the S. cerevisiae population grows successfully when the initial culture is set up.
Show the mark scheme Mark scheme: aseptic technique; provide nutrients or respiratory substrates; a suitable temperature; a pH buffer; stirring or shaking. 'Keep temperature constant' or 'control pH' are ignored.

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From the examiners' report: Most students scored both marks, usually for aseptic technique and a suitable temperature or pH. Saying 'control' the temperature or pH wasn't enough: it has to be suitable for the yeast. Sterilising the culture itself got nothing.

Question 3

A level Paper 2 · June 2018 · Q19(c)(ii) · 2 marks · Medium

Explain the purpose of a flask of nutrient broth with no bacteria added.

Information from earlier in this question:

Earlier part of the same exam question, giving the information this part uses. Part (c)(ii): no bacteria were ever detected in the flask inoculated with nutrient broth that contained no bacteria; explain the purpose of this flask.
Show the mark scheme Mark scheme: it is a control, to check for contamination; 'control group', 'control variable' or 'control condition' are not credited.

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From the examiners' report: Most students said it was a control, but some mixed this up with a control variable or control group. The context points to contamination, so generic answers about comparison got nothing.

Question 4

A level Paper 2 · June 2018 · Q19(c)(iii) · 1 mark · Medium

Suggest why the students should not grow bacteria at 35 °C.

Part (c)(iii): the teacher told the students not to investigate the growth of bacteria at 35 °C; suggest why.
Show the mark scheme Mark scheme: it could encourage the growth of human pathogens; references to denaturation are not credited.

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From the examiners' report: Only about a third of students scored. Wrong answers said the enzymes would denature, which is unlikely at 35 °C, or that growth would be rapid and uncontrollable. The point is that 35 °C is close to body temperature, so human pathogens could grow.

Question 5

A level Paper 2 · June 2022 · Q17(a)(ii) · 2 marks · Hard

List two precautions that keep the preparation of a bacterial culture aseptic.

Information from earlier in this question:

Earlier part of the same exam question, giving the information this part uses. Part (a)(ii): the students prepared the culture by adding a bacterial suspension to a flask of nutrient broth; list two precautions to make the procedure aseptic.
Show the mark scheme Mark scheme: sterile or autoclaved flask, pipette or broth; stopper the flask; disinfect surfaces; work near a Bunsen flame for the upward air flow; flaming the neck of the flask is allowed. An airtight seal is not credited.

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From the examiners' report: Students who used the context sterilised the flasks, broth and pipette and stoppered or flamed the flask. Answers about streaking an agar plate didn't fit the question. The Bunsen flame creates an upward air current; it doesn't kill all the microbes.

Controlling a growth investigation

Question 6

A level Paper 2 · June 2022 · Q17(b)(i) · 1 mark · Medium

Suggest why the flasks were shaken before samples were removed.

Part (b)(i): samples are serially diluted by adding 0.1 cm³ to 9.9 cm³ of water, cells in 0.01 cm³ are counted with a microscope and 0.1 cm³ is spread on agar; suggest why the flasks were shaken first.
Show the mark scheme Mark scheme: so the bacterial cells are evenly distributed.

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From the examiners' report: About half of students scored. Many said the sample needed mixing or that bacteria settle, but missed the key idea: the bacteria must be evenly distributed so the sample is representative.

Question 7

A level Paper 2 · June 2018 · Q19(a) · 1 mark · Medium

State one further variable to control when comparing bacterial growth at two temperatures.

Question 19: three flasks of nutrient broth, two inoculated with 1 cm³ of Bacillus subtilis broth at 20 °C and 30 °C and one with sterile broth at 30 °C, sampled over 3 days for viable bacteria; state one further variable to control.
Show the mark scheme Mark scheme: volume of broth, pH of broth, oxygen concentration, starting number of bacteria, volume removed, or standard mixing before sampling; 'amount' is ignored.

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From the examiners' report: About half of students scored. 'Amount' got no credit as usual, and nor did irrelevant answers such as the size of the flask or light, or describing a control.

Serial dilutions and colony counts

Question 8

A level Paper 2 · June 2018 · Q19(b)(i) · 2 marks · Medium

Use the colonies from Tube F to calculate the viable bacteria in the original sample, in standard form.

Table 19.1 of a serial dilution from Tube A (0.1 cm³ in 9.9 cm³ sterile water) to Tube F, each step 1 cm³ into 9 cm³, and Fig. 19 showing six agar plates: A to C overgrown, D a dense lawn of tiny colonies, E about 40 colonies and F 6 colonies.
Show the mark scheme Mark scheme: 6.0 × 10⁷; maximum 1 mark if not in standard form, and 1 mark for 6 × 10⁶ or 6 × 10⁸.

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From the examiners' report: Fewer than 1 in 5 got both marks. Many were out by a factor of 10 or 100 from the dilutions, and some ignored the instruction to give the answer in standard form.

Question 9

A level Paper 2 · June 2018 · Q19(b)(ii) · 3 marks · Medium

Discuss whether the plate from Tube F was the best one to use for the calculation.

Part (b)(ii): the students disagreed about which tube's result to use; discuss whether the petri dish from Tube F was the most appropriate.
Show the mark scheme Mark scheme: they should have used E, which has more countable colonies, so anomalies have a smaller effect, the sample is more representative and fewer dilutions mean less error; F is countable.

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From the examiners' report: Many students thought F was best, which limited them to 1 mark, and others wouldn't clearly say E. Use the word 'colonies'. Saying someone might miscount E is human error and scored nothing.

Question 10

A level Paper 3 · June 2024 · Q1(b)(i) · 2 marks · Medium

Describe two improvements, other than new equipment, to make a bacterial population estimate more accurate.

Part (b)(i): a method that takes 10 cm³ from a 50 cm³ E. coli culture, makes four 10-fold serial dilutions of 1 cm³ into 9 cm³, spreads 1 cm³ of the final dilution on an agar plate, incubates for 24 h and counts colonies.
Show the mark scheme Mark scheme: mix each dilution; change micropipette tips between transfers; repeat the plating and calculate a mean; photograph the plate to count; or reduce the number of dilutions. 'Repeat the experiment' and general aseptic technique are ignored.

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From the examiners' report: Good answers repeated the plating or reduced the number of transfers. 'Repeat the experiment' wasn't credited, because it suggests starting again with a new population.

Question 11

A level Paper 3 · June 2024 · Q1(b)(ii) · 3 marks · Medium

Calculate the number of E. coli cells in the original 50 cm³ culture from a count of 22 colonies.

Part (b)(ii): the student counts 22 colonies on the agar plate; calculate the total number of E. coli cells in the original 50 cm³ culture, in standard form.
Show the mark scheme Mark scheme: 22 × 10⁵ = 2 200 000, × 5 = 11 000 000, = 1.1 × 10⁷; error carried forward allowed for the working marks.

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From the examiners' report: Many students got this right. Show your working, because marks are given for each stage, and give the final answer in standard form.

Question 12

A level Paper 2 · June 2022 · Q17(b)(iii) · 3 marks · Hard

Describe the calculation steps to estimate the total bacteria in a flask from a microscope count of 52 cells.

Information from earlier in this question:

Earlier part of the same exam question, giving the information this part uses. Earlier part of the same exam question, giving the information this part uses. Part (b)(iii): in one 0.01 cm³ sample the students counted 52 bacterial cells under the microscope; describe the calculation steps to estimate the total number of bacteria in the small flask.
Show the mark scheme Mark scheme: multiply 52 by 1000 to get the number in 10 cm³, multiply by 100ⁿ for n serial dilutions, and multiply by 5 for the 50 cm³ flask; steps can be in any order.

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From the examiners' report: Few students got all 3 marks. The dilution step was the hardest: each 0.1 into 9.9 cm³ dilution is 1 in 100, so multiply by 100ⁿ. Describe the steps as asked, because the number of dilutions isn't given.

Question 13

A level Paper 3 · November 2021 · Q2(d)(iii) · 2 marks · Hard

Describe a laboratory procedure to estimate the size of a bacterial population.

Part (d)(iii) asking for a laboratory procedure the scientists might have used to estimate a bacterial population growing in a closed culture.
Show the mark scheme Mark scheme: serial dilution; grow colonies on an agar plate and count them; scale up the count to estimate the population; flow cytometry is allowed.

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From the examiners' report: Students often couldn't describe a procedure. Few mentioned serial dilution, or scaling up the colony count to the original culture.

Question 14

A level Paper 3 · June 2019 · Q6(b)(i) · 3 marks · Hard

Estimate how many colonies would grow on a plate after a two-step dilution.

Part (b)(i): 20 cm³ of culture contains 1.0 × 10⁶ cells; 5% is made up to 10 cm³ with water, diluted a further ten-fold, and 0.1 cm³ of the final 10 cm³ is spread on an agar plate; estimate the number of colonies.
Show the mark scheme Mark scheme: 50; if wrong, one mark each for 50 000 (5 × 10⁴) and 5000 (5 × 10³).

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From the examiners' report: This multi-step calculation was hard for many, but students who slipped at a later step still picked up working marks for 50 000 or 5000. Write each step down.

Question 15

A level Paper 3 · June 2019 · Q6(b)(ii) · 1 mark · Hard

Explain why an estimate based on four colonies is likely to be inaccurate.

Part (b)(ii): a different serial dilution gave four colonies on an agar plate; explain why the population estimate is likely to be inaccurate.
Show the mark scheme Mark scheme: with so few colonies, small random differences in plating produce large errors when the count is scaled up.

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From the examiners' report: Very few students saw that a low count magnifies error when it is scaled up. Some knew there had been too many dilutions but couldn't express the idea clearly.

Where students lose marks (from OCR examiners' reports and mark schemes)

Each point below is followed by wording that gained marks.

  • Suitable, not just controlled: Students said the temperature or pH of a culture should be 'controlled' or 'kept constant', which wasn't credited, and some suggested sterilising the culture itself.

    ✔ Gains credit: "(Incubate at) suitable temperature" or "maintain optimum pH / use (pH) buffer".

  • A control checks for contamination: Students confused the sterile broth flask with a control variable or control group, or gave generic answers about comparison.

    ✔ Gains credit: "Control; idea of checking for contamination".

  • Pathogens, not denaturation: Asked why bacteria shouldn't be grown at 35 °C, students said enzymes would denature or growth would be uncontrollable.

    ✔ Gains credit: "Idea that it could encourage the growth of human pathogens".

  • The flame makes an updraught: Many thought the Bunsen burner kills all the microbes, and some described streaking an agar plate when the question was about broth.

    ✔ Gains credit: "(Nearby) Bunsen flame (to create upward air flow)"; "use sterile / autoclaved flask / pipette / broth".

  • Evenly distributed: Students said the culture was shaken so it was mixed, or because the bacteria settle, without the reason that matters.

    ✔ Gains credit: "Idea of so bacterial cells are evenly distributed".

  • Pick the plate with most countable colonies: Many students chose the plate with the fewest colonies, or rejected a busier plate because someone might miscount.

    ✔ Gains credit: "Should have used E; (has) more (countable) colonies; anomalies will have smaller effect".

  • Track every dilution factor: Colony count answers were often out by a factor of 10 or 100, and few could deal with several serial dilutions.

    ✔ Gains credit: "Multiply by 100ⁿ (where n is the number of serial dilutions)", then "multiply by 5" to scale up to the 50 cm³ flask.

  • Repeat the plating, not the experiment: 'Repeat the experiment' wasn't credited as an improvement, because it means starting again with a new population.

    ✔ Gains credit: "Repeat the plating (at least three times) and calculate a mean".

Download the questions

Download the PDF pack (15 questions, mark schemes at the end)

Keep practising

Questions, mark schemes and examiners' report extracts © OCR.

Tom Whitburn
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