User:Kathryn Muratore/Notebook/AU CHEM-570 lab prep/2011/08/02

From OpenWetWare
Jump to: navigation, search
Owwnotebook icon.png AU CHEM-570 Lab Prep <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html>      </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>

Objective

  • No growth on yesterday's plates → replate remainder this afternoon
  • DpnI digest of +His/(-)control QuikChange of pMXB10 from yesterday; subsequently transform these samples
  • Run gel with QC samples
  • Double-digest of pTXB1 with BamHI and EcoRI (retrial with higher [DNA])


Bench work

  1. Ran 1.2% agarose gel with QC reactions
  2. Added 1μL of DpnI to each of the QC reactions from yesterday
  3. Double-digest of pTXB1 with BamHI and EcoRI
    • NEB suggests NEBuffer EcoRI for this double-digest, but NEBuffer 3 was used instead.
    • 50μL volume: 10μL 200μg/mL pTXB1, 5μL NEB 3 (10X), 5μL EcoRI (2kU/mL), 5μL BamHI (20kU/mL), 24.5μL dH2O, 0.5μL BSA (100X)
    • 2 hours @ 37°C

Results

Gel

+HispMXB10andcontrolgel08.2.2011labeled.jpg