User:Mariana Ruiz Velasco L./Notebook/IGEM 2010/Wet lab journal/2010/08/30

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So happy I could die!!!!


  • Today, Melvin started doing the LRE transformation described last Thursday (as ligations were left the whole weekend).


  • 1: LRE (XBA I/PST I) with J23100 (ampicilin).
  • 2: LRE (ECO RI/PST I) with plasmid 17 labeled as LRE + p17 Mar and date (chloramphenicol).
  • 3: Control of J23100 (ampicilin).
  • 4: Control of plasmid 17 (chloramphenicol).
  • 5: Control of transformation (BBa_I51020) (ampicilin).
  • 6: Control of competent cells.


Primer's for Click Beetle Luciferase


  • As Dr. Chris Wood gave us two vectors containing luciferases from click beetle (Chroma-Luc™ Reporter Vectors CBRluc and CBG99luc, Promega Corporation), but which don't have the registry's standard, I did primers to add the preffix and RBS, and the suffix. This will help me to work with them.


  • Forward primer 5' -> 3':


GAA TTC GCG GCC GCT TCT AGA GAT TAA AGA GGA GAA AAT GGT GAA GCG TGA GAA AAA TG

Where bold letters indicate the preffix, italics indicate a strong RBS, and normal letters indicate the beginning of the sequence.

  • Reverse primer 5' -> 3':


CTG CAG CGG CCG CTA CTA GTA TTA TTA ACC GCC GGC CTT CTC

Where bold letters indicate the suffix and normal letters indicate the beginning of the sequence.

  • To continue proving the luciferase, I made a restriction for a total of 30μl as follows for both luciferases.


-H2O -------> 8μl

-Buffer 2 --> 3μl

-BSA ------> 1μl

-DNA -------> 15μl

-SPE I -----> 1.5μl

-PST I ------> 1.5μl


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