Melbourne/Lab GV Notebook
From 2007.igem.org
< Melbourne
Revision as of 07:59, 25 October 2007 by PhillipDodson (Talk | contribs)
<Return to Lab notebook> <team home page>
- Synopsis:
The polycistronic gas vesicle gene was obtained in an expression plasmid. We followed the normal process for converting it to a biobrick. The only complicating factor should have been the large size of the part (11Kbp approx). To help with this we used the stratagene XL Quickchange kit for site dirrected mutagenesis, and XL-10 gold ultra competent cells. Things were going swimmingly until we tried to digest the biobrick prefix & suffix and ligate it into BBa_J61035 which carries an RBS with agentamycin handle. The digestion, purification, and ligation processes present a large number of options but very few points of intermediate observation.
Steps:
- Create Glycerol stocks of gas vesicle plasmids and cells supplied by Maura Cannon
- Site dirrected mutagenesis:
- PCR based insertion of Biobrick prefix and suffix, digestion and ligation to BBa_J61035
- Testing Floatation
- Sequencing
- Future directions
