Team:Paris/August 13

Sequencing Minipreps we did yesterday

 * O18 has been diluted to have a concentration of 5µM
 * In each tube : 1µL of O18 diluted
 * Pure water qsp 15µL

Minipreps: Plasmid extraction

 * Protocol (see # 3)  Experiments done by QIAcube


 * List of Minipreps

Glycerol Stocks

 * Protocol (see #2)


 * List of Stocks

Assay to purify a PCR product using the Qiaquick Gel Extraction kit
=> see Gel 1, well n° 8
 * sample used: yesterday PCR product of L130.8 (~40 µL)
 * protocol used: Qiagen PCR purification kit
 * use of buffer QG (Gel Extraction kit) instead of buffer PBI (PCR purification kit)
 * elution in 30 µL of buffer EB
 * 30 µL of purified product + 12 µL of loading blue
 * electrophoresis, 10 µL loaded

==> results: PCR products can be purified using the Qiaquick Gel Extraction kit. We just have to replace the buffer PBI by the buffer QG!

Electrophoresis Setting
4 more transformants of L130 (pFlhB into J61002) are screened by PCR.
 * PCR screening programm; elogation time: 1 min 30
 * template: colonies from the transformation plate
 * positive control: S142 (J61002)
 * negative control: no template
 * primers: O18 and O19

Results of Electrophoresis



 * 1% agarose gel
 * 10 µL loaded

==> The L130 transformants analysed are not correct.

Transformation of the ligation we did yesterday
We transformed L 139, L140, L141 and L142 following the standard protocol using Invitrogen's TOP 10 chemically competent cells. The positive control is a transformation with pUC19 and the negative control has no plasmid.