Team:Hawaii/Notebook/2008-09- 6

From 2008.igem.org

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Things we did today

Wetlab work

Construction of B_H_R Vector

Margaret
  • Yesterday's Restriction Digest
  • Plac+B0030 and Plac+B0034: ethanol precipitation [[1]]
  • pSB1A3, pSB3K3, J33207: de-phosphorylated using SAP
  • Gel Quantification
Gel purified quantification of restriction digest.
  • Some of these products did not appear on the gel, so other methods will be used to generate them.
Gel quantification of the Plac+B0030 and Plac+B0034.
  • Plac+B0030 and Plac+B0034 did not show up on the gel, so the ligation is being performed tonight, pSB1A3 and pSB3K3 plasmid preps from 9/4 showed a good band.
  • Ligation 1:1, followed by Restriction digest with E,S,& P (to cut up any polymers formed and to leave E & S overhang):
  • Plac + B0030 (500ng)
  • Plac + B0030 (100ng)
  • Plac + B0034 (500ng)
  • Plac + B0030 (100ng)
  • pSB3K3 restriction digested with E, P

OriT test

Margaret
  • The biobrick form of OriTr did not show up on the above mentioned gel, so I am:
  • PCR amplifying it from the biobrick paper
  • Extracting DNA from the biobrick registry (1004, 10D) and transforming into DH5-alpha cells. The negative control is pSB1A3-dephosphorylated, the positive control is pSB1A3, from a plasmid prep.

Culturing

Margaret
  • DH5-alpha (for making competent cells)
  • rep, J33207, pSB3K3, and omega interposon

Discussion

Quote of the Day

History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson


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