Plasmid Minipreps (no column)

This is particularly useful for low-copy number plasmids, e.g. pBR322, or cosmids, which often give low yield from typical Qiagen columns. Adapted from the original 1979 alkaline lysis method by Birnboim and Doly.

Note: the quality of the DNA is sufficient for cloning/sequencing purposes, but can have a fair bit of mRNA contamination, and is probably not of transfection quality.

  1. Grow bacterial culture overnight in 5 ml LB + antibiotic.
  2. Sediment bacteria, remove media.
  3. Resuspend bacteria in 250 µL Buffer P1
  4. Add 250 µL Buffer P2
  5. Mix by inverting tube ~5 times.
  6. Let stand at room temperature 2-5 minutes.
  7. Add 250 µL Buffer P3
  8. Mix by inverting tube ~5 times.
  9. Sediment in microfuge at max speed for 10 minutes.
  10. Remove supernatant, add to new tube.
  11. Isoproponal precipitate DNA (without adding additional sodium acetate).
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