Team:Stanford/NotebookPage/9 July 2010

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Meeting Minutes

[http://docs.google.com/present/edit?id=0Ae31et9w_tAhZGZyc3R2ejJfOGNnY2drMmZz&hl=en&authkey=CPHGzfIB Weekly Leader Presentation: Karina]

Agenda

Karina's Notebook

Today's Goal: Digest GFP, RFP, and Terminators. Will do (GFP + T) and (RFP + T) ligations on monday.

Part #'s:

  • GFP: E0040 (cut at S and P)
  • RFP: E1010 (cut at S and P)
  • Term: B1006 (cut at X and P)

Recipe

  • 12.0 uL DNA
  • 1.0 uL each enzyme
  • 5.0 uL NEB buffer 2
  • 5 uL BSA (10x)
  • Sterile H20 to fill up to 50 uL

Mix and put 37º waterbath for 2 or more hours.

Making Gel
To make 50 mL of 0.8% agarose gel, add:

  • .4 g agarose
  • 50 mL TAE (1x)
  • 10 uL EtBr

Loading the Gel

  • add 10 uL loading dye (6x) to digests
  • add 40 uL of dye + digests to wells
  • don't forget to include wells with controls! (uncut plasmids)
  • load 10 uL ladder
  • run at 95 V for about an hour

!!note sizes of plasmids/inserts

GFP plasmid (pSB1A2) --> 2079 bp
GFP insert --> 720 bp
RFP plasmid (pSB2K3) --> 4425 bp
RFP insert --> 681 bp