IGEM:Harvard/2006/DNA nanostructures/Notebook/2006-8-22: Difference between revisions

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Realized that lid oligos were accidentally added in folding of c5.0.A on [[IGEM:Harvard/2006/DNA_nanostructures/Notebook/2006-8-20|8-20]]. Refolding c5.0.A to be just barrel oligos. Will use in PEG fractionation.
Realized that lid oligos were accidentally added in folding of c5.0.A on [[IGEM:Harvard/2006/DNA_nanostructures/Notebook/2006-8-20|8-20]]. Refolding c5.0.A to be just barrel oligos. Will use in PEG fractionation.


=Totals=
 
{| {{table}}
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Working Stocks'''
| align="center" style="background:#f0f0f0;"|'''Working Stocks'''

Revision as of 11:09, 22 August 2006

Folding

Design 5

Realized that lid oligos were accidentally added in folding of c5.0.A on 8-20. Refolding c5.0.A to be just barrel oligos. Will use in PEG fractionation.


Working Stocks Description Pre-Working Stocks Water Total
c5.0A no latches, no aptamers, no lids 1 (86 μL), 4 (2.5 μL), 5 (2.5 μL), 6 (1.5 μL), 7 (1.5 μL) 106 μL 200 μL

Streptavidin Depletion Assay

  • Protocol:
    • Testing Conditions:
      • (A) Unbeaded
      • (B) Beaded
      • (C) Free-streptavidin incubated
      • (D) Free-streptavidin incubated, then beaded
    • Test Solutions:
      • (1) c5.0.A (lidless barrel, no biotinylation)
      • (2) c5.0.Eb (lidless, outside biotinylation)
      • (3) c5.0.Fb (lidless, inside biotinylation)
    • Mix:
      • (B) Beaded
28uL agarose streptavidin beads
40uL of each test solution rxn

Spin down; remove supernatant to new tube.

Load 40uL of the 68uL in gel
      • (C) Free-streptavidin incubated
        • Free-streptavidin concentration needed
          • 1680pmoles on biotinylated boxes + 36pmoles of free biotinylated oligos = 1716pmoles = 1.7nmoles
          • free streptavidin @ 2uM = 2nmol/uL
          • thus, for safety, include 2uL of streptavidin
2uL of 2uM streptavidin
40uL of test solution
26uL of H2O

Load 40uL of the 68uL in gel
      • (D) Free-streptavidin incubated, then beaded
2uL of 2uM streptavidin
40uL of test solution

Wait 5 minutes

Add 28uL of agarose streptavidin beads

Spin down; remove supernatant to new tube.

Load 40uL of the 68uL in gel
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