Preparation of PEI Stocks

Revision as of 16:16, 8 December 2017 by Davebridges (Talk | contribs) (Added notes about testing transfection efficiency of PEI batches.)

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Revision as of 16:16, 8 December 2017 by Davebridges (Talk | contribs) (Added notes about testing transfection efficiency of PEI batches.)

(diff) ← Older revision | Latest revision (diff) | Newer revision → (diff)

Materials

  • Polyethylenimine 25kDa; from Sigma catalog number 764965-1G made at 1 mg/mL

Preparation

  • Dissolve 100 mg in 100 mL sterile ddH2O
  • Stir while slowly adding HCl to pH 7.0
  • Mix for 10 minutes and then recheck pH
  • Filter sterilize through 0.22 um filter
  • Aliquot 500 uL to 1000 uL and store in -80C
  • Thawed solution can be stored at 4C for up to 2 months, label a tube when thawed

Optimization

Each batch of PEI should be tested for efficiency by transfecting cells with GFP at a 1:1-1:6 DNA:PEI ratio and checking the number of fluorescent cells 1-2 days after transfection. Follow the protocol in Production of LentiCRISPR Viruses starting after the table and ending before collecting any media.