Team:Warsaw/Calendar-Main/3 June 2008

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1. DNA (PCR products) gel electrophoresis and isolation of DNA from proper bands
2. Digestion of DNA with HindIII and SalI
3. Digestion of pMPM-T5 + AID with HindIII and SalI
4. Clean-up of products of p.2 and p.3 reactions
5. Ligation of p.4 products (pMPM-T5 + AID transcription fusion with AID-T7 RNA-polymerase translation fusion)
6. Transformation of E.coli TOP10 with ligation product
7. Plating transformants on LB+tetracycline
8. Digestion of pMPM-T5 + transcription fusion with EcoRHI and HindIII
9. Use of T4 polymerase for blunting
10. DNA gel electrophoresis
11. Isolation of DNA from proper band
12. Ligation of product p.11 (pMPM-T5 + T7 RNA-polymerase)
13. Transformation of E.coli TOP10 with ligation product
14. Plating transformants on LB + tetracycline