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From 2013.igem.org

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.
6. Heat shock at 42 °C for 30 seconds.
7. Return to ice for 2 minutes.
8. Add 200 ul LB medium and recover the cells by shaking at 37 °C for 30 min (AmpR)
9. Plate out the cells (10 ul) on Amp LB plates as well as a control (untransformed cells)
10. Incubate at 37 °C. Transformants should appear within 12 hrs.

Result: Colonies (around 10)


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”


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