User:Allison K. Alix/Notebook/Thesis Research/2013/04/15: Difference between revisions
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== | ==Objectives== | ||
* Repeat Gel Electrophoresis on same samples as 4/12/13 (Stained in ethidium bromide for too long | * Repeat Gel Electrophoresis on same samples as 4/12/13 (Stained in ethidium bromide for too long | ||
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==Procedures== | ==Procedures== | ||
Part 1: Repeating Gel Electrophoresis | '''Part 1: Repeating Gel Electrophoresis''' | ||
* Use same procedure as [[User:Allison K. Alix/Notebook/Thesis Research/2013/04/11|04/11/2013]] | * Use same procedure as [[User:Allison K. Alix/Notebook/Thesis Research/2013/04/11|04/11/2013]] | ||
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* Prepare samples the same way | * Prepare samples the same way | ||
Part 2: Reacting AuNP with DNA-thiol | '''Part 2: Reacting AuNP with DNA-thiol''' | ||
1. Mix 250μL 2.5μM ThT/727.5nM thiol-DNA synthesized [[User:Allison K. Alix/Notebook/Thesis Research/2013/04/05|04/05/2013]] with 250μL 4% TEA containing 100mM DTT | 1. Mix 250μL 2.5μM ThT/727.5nM thiol-DNA synthesized [[User:Allison K. Alix/Notebook/Thesis Research/2013/04/05|04/05/2013]] with 250μL 4% TEA containing 100mM DTT | ||
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5. React for 10 minutes | 5. React for 10 minutes | ||
6. Centrifuge 4 times, each time re-dispersing in 500 μL 50mM HEPES buffer | |||
7. Save all supernatants and run Absorbance/Fluorescence Measurements on all | |||
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Revision as of 10:17, 15 April 2013
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Objectives
ProceduresPart 1: Repeating Gel Electrophoresis
Part 2: Reacting AuNP with DNA-thiol 1. Mix 250μL 2.5μM ThT/727.5nM thiol-DNA synthesized 04/05/2013 with 250μL 4% TEA containing 100mM DTT
2. React 10 minutes 3. Extract DTT using 4 2mL aliquots of ethyl acetate. DTT partitions into ethyl acetate and DNA partitions into aqueous phase 4. Mix 232.4μL 1.25μM ThT/363.75nM thiol-DNA with 232.4μL 4.85nM AuNP and 35.2μL sodium citrate buffer
363.75nM/74 = 4.85nM 19.4nM (x) = 4.85nM (250μL) x = 62.5μL AuNP in 187.5μL flitered water 5. React for 10 minutes 6. Centrifuge 4 times, each time re-dispersing in 500 μL 50mM HEPES buffer 7. Save all supernatants and run Absorbance/Fluorescence Measurements on all |