IGEM:University of East Anglia (UEA), Norwich, UK/2009/Notebook/NRP-UEA-Norwich iGEM/2013/06/21
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The restriction digests were retrieved from the freezer while a gel was set up. After the gel had ran for 2 hours, the bands of DNA were visible under ultra violet light so the digest had worked. This meant we could go ahead and remove the fragments of interest and isolate the DNA using a gel cleanup system yielding several samples including four large fragments and two extra large fragments of PSB1C3 plasmid DNA and one small fragment of antGPNeo plasmid DNA. The next stage involved ligation reactions of various different combinations of iGEM plasmid and antGP fragments using the isolated DNA fragments from both today and 19th June. The small antGPNeo fragments were ligated with both the large and extra large PSB1C3 fragments seperately. A negative and postive control were set up. For the postive the small fragment and large fragment of PSB1C3 were ligated. The negative control involved single fragments of antGPNeo and PSB1C3 to see if they self ligate. The samples were left to incubate at 4°C overnight.
Fig: Third Analysis of Restriction Digest products, PSB1C3 and AntGPneo Plasmids cut with ECOR1 and pst1. One fragment removed from each top and middle band of lanes 3 and 5. One Fragment Removed from each middle band of lanes 7 and 9. One Fragment removed from the lower band of lane 11.