User:Mariana Ruiz Velasco L./Notebook/IGEM 2010/Wet lab journal/2010/07/06

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Back to the lab


  • Today, I repeated the gel that I couldn't see the other day.





  • The lanes were as follows:


1. Ladder.
2. 1.2 p-s.
3. 1.7 p-s.
4. 1.9 p-s.
5. 2.2 p-s.
6. 2.8 p-s.
7. 3.1 p-s.
8. 3.3 p-s.
9. 3.5 p-s.
10. 3.8 p-s.
11. 3.9 p-s.
12. 4.2 p-s.
13. 4.3 p-s.
14. Ladder.
15. Empty.
16. Ladder.
17. 4.7 p-s.
18. 4.8 p-s.
19. 4.9 p-s.
20. 2.2
21. 2.8
22. 4.2
23. 4.3
24. 4.7
25. 4.8
26. 4.9
27. Co Neg PCR
28. Co Pos PCR (mut-luz PCR)


Me against the gel


  • In order to quantify the ligation, I ran a gel for 40 min at 95V, which is observed next:




  • The lanes were as follows:


1. Ladder

2. Dephosphated plasmid with J23106

3. Dephosphated plasmid 18

4. Restriction of Luciferase

5. Restriction of Mutated luciferase


Kind of getting used to ligations...


  • As I am planning on repeating the experiments, I did the ligations again...


  • 1: luciferase (green) with J23106


-H2O -----------------------> 9μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 2μL
-Insert (luciferase) ---> 6μL
-Ligase --------------------> 1μL

  • 2: luciferase (green) with plasmid 18


-H2O -----------------------> 8μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 3μL
-Insert (luciferase) ---> 6μL
-Ligase --------------------> 1μL

  • 3: luciferase (red) with J23106


-H2O -----------------------> 10μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 2μL
-Insert (luciferase) ---> 5μL
-Ligase --------------------> 1μL

  • 4: luciferase (red) with plasmid 18


-H2O -----------------------> 9μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 3μL
-Insert (luciferase) ---> 5μL
-Ligase --------------------> 1μL

  • 5: Control (J23106)


-H2O -----------------------> 15μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 2μL
-Ligase --------------------> 1μL

  • 6: Control (plasmid 18)


-H2O -----------------------> 14μL
-Buffer ligase -----------> 2μL
-Vector (plasmid) -----> 3μL
-Ligase --------------------> 1μL

  • The reactions were left at 16°C ON.



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