Team:UCL London/Ligation

From 2010.igem.org

(Difference between revisions)
(New page: {{:Team:UCL_London/templates/v2/headerFullWidth}} ='''Ligation'''= ===Materials=== Plasmid DNA: 15 µL Restriction Enzymes (depend on the requirements of the parts) EcoRI:...)
m (UCL-Ligation moved to Team:UCL London/Ligation: Moved into team namespace)
 

Latest revision as of 20:47, 24 October 2010

UCL IGEM 2010

RETURN TO IGEM 2010

Ligation

Materials

  Plasmid DNA: 15 µL
  Restriction Enzymes (depend on the requirements of the parts)
  
  EcoRI: 1 µL
  PstI: 1 µL
  SpeI: 1 µL
  XbaI: 1 µL
  10× NE buffer: 1.5 µL/1 µL (lid)
  BSA: 1 µL (lid)
  Quick Ligase
  2× Quick Ligation Buffer

Method

1. Digest the DNA as in Restriction Enzyme Digestion Protocol.

2. Heat inactive the restriction enzymes.

3. Run a diagnostic gel before the ligation.

4. After the gel is confirmed, add 1 µL quick ligase, 10 µL ligation buffer, 3 µL backbone and 6 µL inserts (or other volumes depend on the concentration of DNA) into an eppendorf.

5. Leave for 5 minutes (or up to 30 minutes) at room temperature (25°C).

6. Transform the ligated DNA onto competent cells as described in Transformation, method 2 step 4 to 10.


Retrieved from "http://2010.igem.org/Team:UCL_London/Ligation"