User:Mariana Ruiz Velasco L./Notebook/IGEM 2010/Wet lab journal/2010/10/18

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Time is running out!

  • Today, I extracted plasmid from the green click bettle luciferase culture and I left a restriction for 20μl to see if the plasmid contains the luciferase. I also made a restriction with SPEI and PST I to be able to ligate it with LRE restricted with XBAI and PST I.

  • I also made some plates with Cm 30 and Am 30 to do the cotransformation.

  • In the afternoon, I ran a gel to check if the restriction went ok, the gel is shown next:

LRE&lucs 18Oct10.JPG

  • The lanes were as follows:

1. Ladder.

2. Restricted GCB luciferase with ECO RI and PST I.

3. Restricted GCB luciferase with SPEI and PST I.

4. Restricted LRE with XBAI and PST I.

  • Unfortunately, I keep observing these awful dots in the CB luciferases, that is why I ran a gel of both, the plasmid purification and the respective restriction with ECO RI and PST I to try to infer what is happening, the gel is shown next:

CBlucs ppvsrest 18Oct10.JPG

  • The lanes from right to left are as follows:

1, 2. Ladder

3, 5. Plasmid restriction

4, 6. Plasmid purification