Week 5

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Revision as of 10:32, 28 August 2007

07/30/07

• We control bacterial grew (link).
• Transformation for [http://partsregistry.org/Part:BBa_C0012 |C0012] and [http://partsregistry.org/Part:BBa_J52034 |J52034] and [http://partsregistry.org/Part:BBa_B0015 |B0015].
• We get along with the ligation [http://partsregistry.org/Part:BBa_R0051 |R0051]+[http://partsregistry.org/Part:BBa_I13507 |I13507] [http://partsregistry.org/Part:BBa_I763007 |(I763007)] (vector 2 ul, insert 6 ul) and the ligation J04500+ C0051 (I763005) (vector 2 ul, insert 10 ul).

07/31/07

• There isn’t any colony for C0012, so we inoculate I52034 and B0015 colony in 5 ml.
• We strake on plates 30/07 ligations.
• We get along with the J04431 fluorescence test:

-From the glycerol stock we sprout 1 ml at 37°C for 1 hour in agitation.

-We verify the OD and the fluorescence every 15 minutes. The fluorescence compare at the istance t=210min and at the OD=0.5.

• At the end of this test we decide to repeat it to see if we can switch off the fluorescent cells with a major quantity of glucose to confirm our assumptions.

We will attend this test on 07/08.

08/01/07

• We get along with:

-Miniprep of I52034 and B0015;

-Digestion of B0015 with Xba/Pst1;

-Digestion of I52034 with Spe/Pst1;

-Extraction from gel (we can’t see B0015);

-R0015+I13507 (I763007) ligation which doesn’t give colonies so we transform with the other 10 ul of ligation;

-J04500+C0051 (I763005) ligation which gives colonies so we inoculate in 5ml;

-Transformation for C0012 (4ul) and for J22101 (2ul);

-A new R0051+ I13507 (I763007) (vector 4ul, insert 8ul) ligation.

08/02/07

• Today we do:

-Miniprep of J04500+C0051 (I763005);

-Control digestion of 5 ul with Eco/Pst1;

-A new B0015 digestion with Xba/Pst1;

-Digested run on electroforesi gel but we have problems with gel because we don’t see the band;

-Band exstraction for B0015 e I763005;

-We observe thre aren’t colonies for C0012, but there are colonies for R0051 + I13507 (I763007), so we identify the correct ligation protocol;

- R0010, P0412, C0012 transformation;

-Inoculation in 5 ml for R0051 + I13507 and for J22101;

- R0051 + I13507 (I763007) fluorescence test to verify if the filter for RFP is adapted, but it isn’t.

08/03/07

• We get along with:

-Miniprep for J22101 and for R0051+ I13507 (I763007);

- J22101 digestion with Xba/Pst1 ;

- Control digestion of R0051 + I13507 (I763007);

- J04500+C0051 (I763005) and B0015 and J22101 and R0051 + I13507 (I763007) run on electroforesi gel and J04500+C0051 (I763005) and J22101 estraction;

-Glycerol stock preparation;

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