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From 2011.igem.org

Tuesday

Looked at our plates from yesterday, we've got colonies on all of them, yaii - and they are white! But our fungi have not grown at all, to bad, but we will try again today.

Labwork

• User PCR on PSB1C3

• Grow cells: UserB1 and ControlBL21. OD measurements show that the UserB1 cells are not growing, hmm, we'll do it again tomorrow.

• Miniprep on 2C9 and UserB1. The Nanodrop measurements are very low, wonder what must have happened. The UserB1 miniprep was for sequencing - this was skipped, because we realized that the vector that the UserB1 not is the same as the expression vector we are using, thus we didn't have the sequence to make the primers.

• O/N digestions are purified by using the PCR purification kit, but we will not continue with these, since our plates with the digestions from yesterday looks fine.

• User PCR on Expr. vector, 2C9 (1-807), 2C9 (808-1434).

• Digestion of UserPsB1C3 with Pac1 and Nt.BbvCI (2 h by 37 degrees)

• User mixture with UserEnzyme and the PCR products: UserExpr. vector, User2C9(1-807), User2C9(808-1434) and Digested UserPSB1C3. The Mixture incubates 15 min by 37 degrees and 15 min by roomtempareture.

• Transformation of the UserMixture to XLblue cells on a CAM/IPTG plate.

• Colony PCR on the A2 plates, to check for insert. Made O/N cultures and plated out at the same time.

• O/N cultures of the ControlBL21 and the UserB1 - this time 4 colonies are picked.

• Made amp plates

• Checked the TLC from friday again

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