User:Madeleine Y. Bee/Notebook/Single Molecule Fluorescence/2013/01/29: Difference between revisions
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== | ==January 29, 2013== | ||
* | *Concentration calculations- | ||
1) make 100μL of 1μM 4 strand DNA:<br> | |||
2.656μL 4 strand DNA in 97.344μL filtered H<sub>2</sub>O <br> | |||
2) make 100μL of 1μM 8 strand DNA:<br> | |||
1.428μL 8 strand DNA in 98.527μL filtered H<sub>2</sub>O <br> | |||
3) make 6.49nM Molecular Beacon: <br> | |||
731nM(V)=6.49nM(1000μL) <br> | |||
V=8.878μL MB in 991.12175μL unfiltered H<sub>2</sub>O <br> | |||
4) make TRIS buffer (pH of 7.2): <br> | |||
(121.14g/mol TRIS)(5mmol/L)(1mol/1000mmol)(50mL)(1L/1000mL)=0.030285g TRIS in 50mL solution <br> | |||
*combine 0.0352g TRIS, 9 drops HCl, 5mL distilled H<sub>2</sub>O, 0.0025g TRIS, 20mL H<sub>2</sub>O <br> | |||
*result: 5mM TRIS buffer solution with a pH of 7.2 <br> | |||
5) make 3.5μM ThT <br> | |||
(318.86g/mol ThT)(3.5mmol/L)(1mol/10^3mmol)=1.116g/L(1L/1000mL)(10mL)=0.01116g ThT in 10mL solution <br> | |||
*dissolve 0.0134g ThT (massed value, approximated for 0.01116g) in 10mL filtered H<sub>2</sub>O : 3.5mM ThT solution <br> | |||
*dilute 10μL 3.5mM ThT solution in 9990μL filtered H<sub>2</sub>O : 3.5μM ThT solution <br> | |||
6) make DNA solutions at varying concentrations: <br> | |||
DNA (4 strand) concentration: 265.6μM <br> | |||
DNA (8 strand) concentration: 142.8μM <br> | |||
*1:265.6(V)=3.5(250μL) <br> | |||
V=3.29μL 4 strand DNA in 246.71μL TRIS buffer solution <br> | |||
*2:2.82μL 4 strand DNA in 247.18μL TRIS buffer solution <br> | |||
*3:2.35μL 4 strand DNA in 247.65μL TRIS buffer solution <br> | |||
*4:1.88μL 4 strand DNA in 248.12μL TRIS buffer solution <br> | |||
*5:1.41μL 4 strand DNA in 248.59μL TRIS buffer solution <br> | |||
*6:142.8(V)=3.5(250μL)<br> | |||
V=6.13μL 8 strand DNA in 243.87μL TRIS buffer solution <br> | |||
*7:5.25μL 8 strand DNA in 244.75μL TRIS buffer solution <br> | |||
*8:4.38μL 8 strand DNA in 245.62μL TRIS buffer solution <br> | |||
*9:3.50μL 8 strand DNA in 246.50μL TRIS buffer solution <br> | |||
*10:2.63μL 8 strand DNA in 247.38μL TRIS buffer solution <br> | |||
7)mix 100μL of each of the 10 DNA solutions with 100μL 3.5μM ThT | |||
Revision as of 08:45, 2 April 2013
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January 29, 2013
1) make 100μL of 1μM 4 strand DNA:
5) make 3.5μM ThT
6) make DNA solutions at varying concentrations:
V=3.29μL 4 strand DNA in 246.71μL TRIS buffer solution
V=6.13μL 8 strand DNA in 243.87μL TRIS buffer solution
7)mix 100μL of each of the 10 DNA solutions with 100μL 3.5μM ThT
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