Notebook : July 8

Summary:

FNR regulator system:

• performed a digestion for AmilCP and LacZ with Not I, Xho I, EcoR I and PST I.
• electrophoresis for those digest products
• enrichment culture for clone of the briobrick BBa_K1155000
• electrophoresis for extracted products of last friday

Lab work

Restriction digest

For some of the enzymes which are possible to use one common buffer, we did double digest DNA enzyme buffer AmilCP1 Not I Orange AmilCP2 Not I Orange AmilCP1 Xho I Red AmilCP2 Xho I Red AmilCP1 EcoR I Orange AmilCP2 EcoR I Orange LacZ1 EcoR I + PST I Orange LacZ2 EcoR I + PST I Orange Volume added: DNA 2(µl) buffer 2(µl) Enzyme(foreach) 0.5(µl) H20 15.5(µl) total 20(µl) Notes: by using double digest, an informatics tool available in Fermentas web site, it is possible to use one buffer for both EcoR I and PST I. 2. The electrophoresis (135 V for 30 minutes) for digest products were migrated in agarose gel (0.5X). Volume added: for each well plasmid 10µl OR Pfnr 5µl Blue Results: 3. Overnight incubation at 37°C with agitation. 4. The electrophoresis for the plasmids which we had extracted last week Volume added: Pfnr2 R1P R2 A1P 10µl 10µl 10µl 10µl Migration at 100V for 25mins then continued till 45 minutes: Results: The concentration of Pfnr2 was detected by NanoDrop : C = 1788.8 ng/µl 260/280 = 2.08 We considered that our extraction of plasmid DNA was not successful, we need to perform another extraction DNA.