cDNA phage stock and in vivo excision

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cDNA phage stock and in vivo excision


Phage stock

  1. Start an overnight of XL1-Blue from a fresh plate in LB (no antibiotics)
  2. Add 100 l XL1-Blue o/n to 3 ml of lambda top agarose at 50°C
  3. Vortex for 10 sec
  4. Quickly pour onto pre-warmed LB plates, tilt plate to spread evenly
  5. Spot 1-3 l (usually use 2 l) of phage stock onto plate using pipettor; can do 4-6 cDNAs on one plate if careful
  6. Incubate o/n at 37°C to get large plaque
  7. Transfer plaque using big end of pasteur pipet to 0.5 ml SM in micro c/f tube
  8. Add one drop of chloroform
  9. Cap well and vortex
  10. Store at 4°C, do not freeze

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