Cosmid prep or scaled up mini prep

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1. Grow 3-4 tubes of each cosmid in 5 ml media overnight

2. Harvest a total of 4 ml of each tube separately in 2 ml centrifuge tubes

3. Resuspend completely in 200 l Solution I*

4. Add 400 l Solution II* and invert until solution clears

5. Add 300 l Solution III* and invert gently

6. Hold on ice for 10 min

7. Spin 10 min at 4°C and transfer s/n to new tube

8. Phenol/chloro/IAA one time with equal volume

9. Spin 1 min at 4°C and transfer upper layer to new tube

10. Ppt with 0.8 vol isopropanol

11. Spin 20 min at 4°C

12. Remove s/n and allow pellet to dry

13. Resuspend in ~ 50 l of TE + RNase

14. Cosmids => Digest 5-10 l with either EcoRI or Hind III

*refer to Alkaline Lysis Solution I, II, or III recipes in card box

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