Preparation of PEI Stocks: Difference between revisions
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wrote instructions for making PEI stocks |
Added notes about testing transfection efficiency of PEI batches. |
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* Aliquot 500 uL to 1000 uL and store in -80C | * 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 | * 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. | |||
[[ Category: CRISPR ]] | |||
[[ Category: Cell Culture ]] | |||
[[ Category: Tissue Culture ]] | |||
[[ Category: Transfection ]] | |||
[[ Category: Molecular Biology ]] | |||
Latest revision as of 16:16, 8 December 2017
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.