Team:Exeter/lab book/novpol/wk6

Operon Construction  |  Glycobase

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**Monday 13/08/12**

Added ligation mix of HAS-terminator, cyclodextran-terminator, ompA-SacB and ompA-cyclodextran to Top10 competent E.coli cells and transformed onto ampicillin plates.

**Tuesday 14/08/12**

Transferred HAS-terminator, cyclodextran-terminator, ompA-SacB and ompA-cyclodextran to liquid broth containing ampicillin.

**Wednesday 15/08/12**

Performed mini-prep of HAS-terminator, cyclodextran-terminator, ompA-SacB and ompA-cyclodextran. Used the nanodrop machine to confirm nucleic acid present. Digested the fragments and ran them on a gel using electrophoresis. All of the genes were single banded or absent, confirming that the ligation had not worked and only plasmid DNA was present.

Used previous digestion mix of HAS, cyclo, SacB and ompA to attempt ligation again using the 3A assembly protocol, into digested tetracycline plasmids. Transformed ligation mix into Top10 competent E.coli cells and plated out on tetracycline plates.

**Thursday 16/08/12**

None of the plates had successfully grown any cultures so stopped lab work for the weekend.