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From 2011.igem.org

Contents 1 Kevin 1.1 Biobrick the YFP:tetR fusion protein 1.1.1 PCR 1.1.2 Following steps 1.1.3 To be continued 2 Adrien

Kevin

Biobrick the YFP:tetR fusion protein

Let's start again with good primers to biobrick the protein.

PCR

3 Temperatures of primer annealing: 85 - 58 - 48 °C
Primers : pFX234 FW - pFX234 Reverse CP

Following steps

PCR Purification, digestion in ES, PCR Purification again, Ligation

To be continued

Transformation, sequencing and glycerol.

Adrien

-80°C PY79 are resistant to TetR up to 75 μg.mL ^-1 which is better than the 150μg.mL ^-1 than the -20°C glycerols of PY79.
Conclusion: to do efficient electro-poration or chemical transformation into Bacillus and using the multi-host vector pHM3 we must use PY79 from the -80°C glycerol.
Tomorrow, Babak will make four glycerol for the -20°C that are going to replace the existing one out of the 4 falcon I launched today.

Cambridge 2008 amylase test was redone with control on the same plate. To be continued by Babak.