IGEM:Harvard/2006/DNA nanostructures/Notebook/2006-8-17

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(2nd PEG Precipitation)
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* conduct 2nd PEG precipitation on yesterday's precipitated samples (5.0A, C, D 7%-10%)
* conduct 2nd PEG precipitation on yesterday's precipitated samples (5.0A, C, D 7%-10%)
* protocol: mix all ingredients, add nanostructures last, total volume is 100 mL, incubate on ice for 15 min, spin for 10 min at 16k rcf, pipet off supernatant, resuspend pellet in 100 {{ul}} water
* protocol: mix all ingredients, add nanostructures last, total volume is 100 mL, incubate on ice for 15 min, spin for 10 min at 16k rcf, pipet off supernatant, resuspend pellet in 100 {{ul}} water
 +
* CONCLUSIONS: we seem to be unable to perform 2 PEG precipitations because all of the nanostructures ended up in the supernatant. we will repeat the 1st PEG precipitation with c5.0 A, C, D at 6% and probably not repeat another PEG if the protection assay is okay.
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==Magnetic Streptavidin Protection, Take 3==
==Magnetic Streptavidin Protection, Take 3==

Revision as of 17:32, 17 August 2006

2nd PEG Precipitation

  • conduct 2nd PEG precipitation on yesterday's precipitated samples (5.0A, C, D 7%-10%)
  • protocol: mix all ingredients, add nanostructures last, total volume is 100 mL, incubate on ice for 15 min, spin for 10 min at 16k rcf, pipet off supernatant, resuspend pellet in 100 μL water
  • CONCLUSIONS: we seem to be unable to perform 2 PEG precipitations because all of the nanostructures ended up in the supernatant. we will repeat the 1st PEG precipitation with c5.0 A, C, D at 6% and probably not repeat another PEG if the protection assay is okay.
Final PEG %PEG μLNaCl μLc5.0 μLdH2O μL
735104015
840104010
94510405
105010400


lane contents loading dye
18 μL untreated 5.0.A3 μL
220 μL 5.0.A.1 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
38 μL 5.0.A.1 in 7% PEG and 0.5 M NaCl (pellet)3 μL
420 μL 5.0.A.2 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
58 μL 5.0.A.2 in 7% PEG and 0.5 M NaCl (pellet)3 μL
620 μL 5.0.A.1 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
78 μL 5.0.A.1 in 8% PEG and 0.5 M NaCl (pellet)3 μL
820 μL 5.0.A.2 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
98 μL 5.0.A.2 in 8% PEG and 0.5 M NaCl (pellet)3 μL
101 kb+ ladder3 μL
112.7 μL p73083 μL
1220 μL 5.0.A.1 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
138 μL 5.0.A.1 in 9% PEG and 0.5 M NaCl (pellet)3 μL
1420 μL 5.0.A.2 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
158 μL 5.0.A.2 in 9% PEG and 0.5 M NaCl (pellet)3 μL
1620 μL 5.0.A.1 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
178 μL 5.0.A.1 in 10% PEG and 0.5 M NaCl (pellet)3 μL
1820 μL 5.0.A.2 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
198 μL 5.0.A.2 in 10% PEG and 0.5 M NaCl (pellet)3 μL
20empty-
218 μL untreated 5.0.C3 μL
2220 μL 5.0.C.1 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
238 μL 5.0.C.1 in 7% PEG and 0.5 M NaCl (pellet)3 μL
2420 μL 5.0.C.2 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
238 μL 5.0.C.2 in 7% PEG and 0.5 M NaCl (pellet)3 μL
2420 μL 5.0.C.1 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
258 μL 5.0.C.1 in 8% PEG and 0.5 M NaCl (pellet)3 μL
2620 μL 5.0.C.2 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
278 μL 5.0.C.2 in 8% PEG and 0.5 M NaCl (pellet)3 μL
2820 μL 5.0.C.1 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
298 μL 5.0.C.1 in 9% PEG and 0.5 M NaCl (pellet)3 μL
301 kb+ ladder3 μL
312.7 μL p73083 μL
3220 μL 5.0.C.2 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
338 μL 5.0.C.2 in 9% PEG and 0.5 M NaCl (pellet)3 μL
3420 μL 5.0.C.1 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
358 μL 5.0.C.1 in 10% PEG and 0.5 M NaCl (pellet)3 μL
3620 μL 5.0.C.2 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
378 μL 5.0.C.2 in 10% PEG and 0.5 M NaCl (pellet)3 μL
38-40empty-
418 μL untreated 5.0.D3 μL
4220 μL 5.0.D.1 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
438 μL 5.0.D.1 in 7% PEG and 0.5 M NaCl (pellet)3 μL
4420 μL 5.0.D.2 in 7% PEG and 0.5 M NaCl (supernatant)3 μL
458 μL 5.0.D.2 in 7% PEG and 0.5 M NaCl (pellet)3 μL
4620 μL 5.0.D.1 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
478 μL 5.0.D.1 in 8% PEG and 0.5 M NaCl (pellet)3 μL
4820 μL 5.0.D.2 in 8% PEG and 0.5 M NaCl (supernatant)3 μL
498 μL 5.0.D.2 in 8% PEG and 0.5 M NaCl (pellet)3 μL
501 kb+ ladder3 μL
512.7 μL p73083 μL
5220 μL 5.0.D.1 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
538 μL 5.0.D.1 in 9% PEG and 0.5 M NaCl (pellet)3 μL
5420 μL 5.0.D.2 in 9% PEG and 0.5 M NaCl (supernatant)3 μL
558 μL 5.0.D.2 in 9% PEG and 0.5 M NaCl (pellet)3 μL
5620 μL 5.0.D.1 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
578 μL 5.0.D.1 in 10% PEG and 0.5 M NaCl (pellet)3 μL
5820 μL 5.0.D.2 in 10% PEG and 0.5 M NaCl (supernatant)3 μL
598 μL 5.0.D.2 in 10% PEG and 0.5 M NaCl (pellet)3 μL
60empty-

Magnetic Streptavidin Protection, Take 3

  • Gel:
Lane Component Amount
11kb ladder10uL
2p7308 (44nM)7.5uL
3c5.0.D wash40uL
4c5.0.D final40uL
5c5.0.8(b) wash40uL
6c5.0.8(b) final40uL
7c5.0.Eb wash40uL
8c5.0.Eb final40uL
9c5.0.Fb wash40uL
10c5.0.Fb final40uL
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