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Team:Groningen/Notebook/Wetwork 20July2012
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Revision as of 18:40, 16 September 2012
Plan for today:
1. plasmid isolation from E.coli Dh5A
2. ligate alsT promoter and GFP into pSB1C3 and pSac-Cm (B. subtilis backbone)
3. E.coli transformation for both
1. Plasmid isolation pSB1C3+insert from iGEM Groningen 2010
E.coli DH5a strong magenta = insert device OK!
plasmid concentration: 129,1 ng/ul
Purification of PCR product using gel DNA extraction kit
diluted in 50ul TE (elution buffer)
concentration: 35,6 ng.ul
Ligation pAlsT-GFP into backbone pSB1C3 (and pSac-Cm done next week 23/07)
Cut parts with restiction enzymes
alsT: EcoRI + SpeI
GFP: XbaI + PstI
backbone: EcoRI + PstI
Digestion reaction and subsequently ligation of:
1) alsT150-GFP-BB
2) alsT250-GFP-BB
3) alsT300-GFP-BB
4) alsT500-GFP-BB
3. Transformation of E.coli DH5a
4x promoter alsT testing devices
1x negative control
= 5x LB plate + 15 ug/ml Cm
Our sponsors:
