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LABORATORY PROCEDURES FOR ANIMAL & HUMAN CELL LINES
REFERENCE NO: AHC/1998/3/18.104.22.168 Appendix 7
TITLE: DISMANTLING OF SOUTHERN BLOT AND FIXATION OF DNA TO MEMBRANE
protocol AHC/1998/3/3.1/2.3 Appendix 6.
Remove membrane and place on whatman 3MM chromatrography paper DNA side up. Put at 37oC for 4-6h.
While membrane is drying, place the used gel in electophoresis buffer + ethidium bromide (1mg/ml) for 30 mins. Place this gel on the transilluminator to check for any residual DNA. Record the results of restaining.
When dry wrap membrane in 'Saran wrap' or 'Cling Film'.
Switch UV transilluminator on for 5min.
Place wrapped membrane (DNA side-down) onto the transilluminator and fix for 4.5-5 minutes.
Seal wrapped membrane in a polythene bag.
The membrane can now be stored in a cool dry place indefinitely.
- Remove metal plate, towels and filter paper squares from the blot.
- Carefully peel off membrane and rinse (DNA-side up) in 3 x SSC using an appropriate dilution of 20 X SSC, see
Records: note any residual staining after restaining the gel and the exact fixation time.
Guidelines prepared for CABRI by CERDIC, DSMZ, ECACC, INRC, November 1998
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Copyright CABRI, 1998
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