Lidstrom:Preparing Freezer Cell Stocks
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- Grow ~ 5 mL of E. Coli overnight with appropriate antibiotics.
- Centrifuge for ~2+ minutes & pour off supernatant.
- Resuspend in 900 uL of HY (hypho) and 100 uL of filter-sterilized DMSO. Hypho is better than LB for this; I hear it buffers.
- must use a filter that won't dissolve in DMSO . Most of the filters we have are not appropriate.
- Store at -80oC.
- Test stock after frozen a while.
Filtering DMSO: don't dissolve the filter
- Pure DMSO dissolves many of the filters common in our lab.
- "To prepare a sterile solution, use a teflon or nylon membrane to sterile-filter the DMSO; do not use a cellulose acetate membrane." Sigma/Source