IGEM:Harvard/2006/vlau/Notebook/2006-7-25

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Pre-Working Stocks & Working Stocks

1. Rxn Mixtures

Description Plate Locations Total
c4.0.2: core top lid oligosbox 4_A: row F (7-12), row G (1-12), row H (1-12); box 4_B: row A (1-3)33
c4.0.3: core bottom lid oligosbox 4_B: row A (4-12), row B (1-12), row C (1-12)33
Working Stocks Description Pre-Working Stocks Water Total
c4.0Fb.corelatch design 1, inside biotin1 (66 μL, 2 (33 μL), 3 (33 μL), 4b (4 μL)64 μL200 μL
c4.0Fb.latcheslatch design 1, inside biotin8 (4 μL), 11 (2 μL), 12 (2 μL)--
c4.0Gb.corelatch design 1, outside biotin1 (66 μL, 2 (33 μL), 3 (33 μL), 6b (4 μL)64 μL200 μL
c4.0Gb.latcheslatch design 1, outside biotin8 (4 μL), 11 (2 μL), 12 (2 μL)--
c4.0Hb.corelatch design 2, inside biotin1 (66 μL, 2 (33 μL), 3 (33 μL), 4b (4 μL)64 μL200 μL
c4.0Hb.latcheslatch design 2, inside biotin9 (4 μL), 13 (2 μL), 14 (2 μL), 23 (4 μL), 24 (2 μL), 25 (2 μL)--
c4.0Ib.corelatch design 2, outside biotin1 (66 μL, 2 (33 μL), 3 (33 μL), 6b (4 μL)64 μL200 μL
c4.0Ib.latcheslatch design 2, oustide biotin9 (4 μL), 13 (2 μL), 14 (2 μL), 23 (4 μL), 24 (2 μL), 25 (2 μL)--


2. Protocol

   - mixed 10 μL of each 50 μM stock oligo to make pre-working stocks
   - mixed listed pre-working stocks to make working stocks where final concentration of each oligo is 250 nM

Folding Rxns w/ Biotinylated Oligos

1. Reagents (for c4.0Fb, c4.0Gb, c4.0Hb, c4.0Ib)

  - 9 μL p7308 scaffold = (10 nM)/(44 nM) * 40 μL
  - 16 μL oligos = (100 nM)/(250 nM) * 40 μL
  - 4 μL 10x folding buffer (500 mM HEPES pH 7.5, 500 mM NaCl, 100 mM MgCl2)
  - 11 μL dH2O
  - total volume: 40 μL

2. Negative Controls

  - 1st (no oligos): 9 μL scaffold, 16 μL dH2O, 4 μL buffer, 11 μL dH2O
  - 2nd (no scaffold): 9 μL dH2O, 16 μL oligos, 4 μL buffer, 11 μL dH2O
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