Team:Groningen/Labwork/1 August 2013

Mirjam

Plates for the transformation of ΔDes, ΔCheY and ΔDes/ΔCheY did not show any colonies. The plates for eYFP showed some colonies on the plates with the transformation to E.coli. The plates with RFP and the ligation of Pdes/Chey did not show any colonies. Also the plates with a transformation of GFP240 and GFP0840 showed colonies.
Inoculated the colonies into LB with ampicillin to perform a miniprep later on.
Made a new restriction digestion for RFP, eYFP and BBa_k823823 + LacI with EcoRI and PstI. Did a gel purification to obtain the purified fragment.
Inoculated B.subtilis colonies for transformation.
Made a restriction for Pdes with EcoRI and BamHI, for CheY with BamHI and PstI and for BBa_k823823 with EcoRI and PstI.

Chaline

Made a miniprep of eYFP and RFP to get a higher concentration.
Did a gel purification to obtain purified fragments of eYFP, RFP and 3BB.

Sebas

Sander

Made a plasmid purification of GFP BBa_E0240 and got a concentration of 21,5 and also did a purification of GFP BBa_E0840 and got a concentration of 13,5.(placed in temp box labeled as GFP 0240 and GFP 0840)

Claudio

The plates prepared yesterday show colonies. 4 colonies per plate are inoculated in LB + Ampicillin in order to isolate the plasmid on the day after and check for the presence of the desired insert.

To the tube, containing the purified BBa_K823023 + lacI digested with XbaI and PstI, 1µl SpeI is added and the tube is incubated for 1h at 37°C. After the treatment the backbone is ligated with BBa_E0840 and BBa_E0240 (already digested and purified yesterday with XbaI and PstI). The two ligation products are transformed into E. Coli DH5α and plated on LB + Ampicillin.

Bacillus Subtilis is inoculated in LB.

Inne & Sander

Did a inoculation of 2 colonies from the delta cheY plates and 2 colonies of the delta des Both plates were made yesterday.

Sample	Strain	Selection marker
1	Delta cheY 1	5 ug/ml cm
2	Delta cheY 2	5 ug/ml
3	cheY negative control	5 ug/ml cm
4	Delta des 1	5 ug/ml cm 5 ug/ml km
5	Delta des 2	5 ug/ml cm 5 ug/ml km
6	Delta des negative control	5 ug/ml cm 5 ug/ml km