Team:Paris Bettencourt/Notebook/Trojan Horse/Tuesday 16th July.html

Trojan Horse

16^th July

Anne with Nico, Marguerite and Yonatan

Transformation of pUC18 (NEB, NEB CC, BL21)

1) Thaw competent cells on ice.

2) Place 20 ul of cells in a pre-chilled Eppendorf tube.

3) add o.5ul of plasmid to the cells

4) Mix gently by flicking the tube.

5) Chill on ice for 10 minutes.

4) Heat shock at 42 °C for 30 seconds.

5) Return to ice for 2 minutes.

6) Add 200 ul LB medium and recover the cells by shaking at 37 °C for 30 min (AmpR)

7) Plate out the cells (10 ul) on Amp LB plates as well as a control (untransformed cells)

8. Incubate at 37 °C. Transformants should appear within 12 hrs.

Result: Colonies (around 10)

Vincent and Aude

In silico cloning: sRNA gBlocks into pUC18 and litmus28-GFP

Successful with both Gibson assembly and regular cloning.

See file: “in silico cloning trojan horse.geneious”

Vincent

streaking Ortiz’s strain ELS-41 and ELS-13 containing Litmus28i_J23115-B0032-GFP and M13K07, respectively.