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Prof. Martin Jinek

Why did we contact them?

We spoke with Prof. Martin Jinek, Professor at the Department of Biochemistry at the University of Zurich and head of the Jinek Laboratory. He has many years of experience working with CRISPR-Cas systems and was part of the original research team, alongside Emmanuelle Charpentier and Jennifer Doudna, that first described how CRISPR-Cas9 can be used for genome editing. We asked him which Cas protein would be most suitable for our project and whether we should purify the protein ourselves or order it commercially.

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Prof. Martin Jinek

Discussion

Protein purification

  • Purifying the proteins ourselves would take about one week with 1–2 people working on it, plus supervision from someone in his lab.
  • Self-purification could yield about 10 mg of protein, which is 2–5 times more than ordering a kit.
  • The proteins would need to be stored at –80 °C

Pros and cons of purification vs. ordering

  • Ordered kits give less protein and are more expensive.
  • Purification requires more effort but produces a higher yield.
  • The decision depends on how much protein is needed for our experiments.

Choice of Cas12a protein

  • Cas proteins usually work best above 37 °C.
  • LbCas12a is probably best suited for room temperature.
  • FnCas12a is commonly used in the Jinek lab.
  • He has some Lb clones stored but there are no side-by-side stability studies.
  • The stability of Cas proteins after lyophilization is unknown and must be tested.

Detection and fluorescent output

  • He has experience with Cas and quencher/fluorescence output systems.
  • High concentrations of fluorescent reporters can be visible by eye, but this depends strongly on the fluorophore. Machines can filter wavelengths more precisely.
  • Reporters can be adapted when ordering.
  • Suggested looking again into colorimetric output systems as an alternative and testing them if time allows.

Main Takeaways

  • Protein purification in his lab is possible and yields more protein than ordered kits.
  • LbCas12a seems most suitable for room temperature applications.
  • We can adapt fluorophores in the reporter system when ordering.
  • Fluorescent output might be visible to the naked eye at high concentrations, but should be confirmed experimentally.

Integration

We implemented his advice by:

  • Planning to purify Cas12a proteins ourselves in his lab.
  • Considering LbCas12a as a detection candidate for our test.
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