SBB11Ntbk-MaryWang

Mary Wang 14:18, 17 February 2011 (EST)

Analytical gel run on part ssb1139: Used 2uL product from previous day and 5uL dye. Analytical gel shows that the part was done incorrectly. Analytical Gel2- lane 5.

REdone (x2)

One: 24uL ddH2O, 3.3uL 10X expand buffer, 3.3 uL dNTP, 1 uL ss45r, 1uL ss45f, 0.5uL template DNA, 0.5 expand polymerase 1

Two: 21.7uL ddH2O, 3.3uL DMSO, 3.3uL 10X expand buffer, 3.3uL dNTP, 1uLss45r, 1uL ss45f, 0.5uL template DNA, 0.5 expand polymerase 1

Run at 2k45 instead of 2k55

Digest pBjh1601KA-Bjh2296 using: 8uL of eluted PCR product, 1uL of NEB Buffer 2, 0.5uL EcoRI and 0.5uL BamHI. Prep Gel run- shown on PrepGel2- lane 4. Cut out ~837 part- melted with 600uL ADB buffer then froze.

Mary Wang 14:16, 15 February 2011 (EST)

PCR ss45r/ss45f on MG1655 genome.

Used Expand polymerase because it was a genome.

Put template DNA into PCR tube first then the Expand polymerase second.

Did not dilute the oligos.

Used cloning by PCR- basic PCR method.

Amounts used: 24uL ddH2O, 3.3uL 10X expand buffer, 3.3uL dNTP, 1uL Oligo 1, 1uL Oligo 2, 0.5 Expand polymerase 1, 0.5uL Template DNA

Run at 2k55