User:Karmella Haynes/Notebook/PcTF Genomics/2013/07/05: Difference between revisions
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| <u>Wells</u> || <u>Plasmid</u> || <u>DNA</u> || <u>Volume + dH<sub>2</sub>O</u> || <u>Opti-MEM</u> || <u>PLUS</u> || <u>Lipo</u> || | | <u>Wells</u> || <u>Plasmid</u> || <u>DNA</u> || <u>Volume + dH<sub>2</sub>O</u> || <u>Opti-MEM</u> || <u>PLUS</u> || <u>Lipo</u> || | ||
|- | |- | ||
| 1 || KAH160/MV1 || 2 μg || | | 1 || KAH160/MV1 || 2 μg || 5.8 μL +14.2 || 570 μL || 2.5 μL || 7.5 μL | ||
|- | |- | ||
| 2 || KAH160/MV1 || 2 μg || | | 2 || KAH160/MV1 || 2 μg || 5.8 μL +14.2 || " || " || " | ||
|- | |- | ||
| 3 || mock || none || 0 μL + 20.0 || " || " || " | | 3 || mock || none || 0 μL + 20.0 || " || " || " | ||
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| 4 || none || none || --- || --- || --- || --- | | 4 || none || none || --- || --- || --- || --- | ||
|} | |} | ||
> Add 570 μL Opti-MEM to each 20 μL DNA sample <br> | |||
> Add 2.5 μL PLUS reagent to each DNA+Opti-MEM sample. 5 min/room temp<br> | |||
> Add 7.5 μL Lipofectamine LTX to each DNA+Opti-MEM+PLUS reagent sample. 30 min/ room temp <br> | |||
> Add 600 μL complexes to each well (final volume of medium = 4 ml/well)<br> | |||
> Grow cells at 37°C overnight<br> | |||
Results (7/06/13) | |||
* Success! Very high levels of RFP expression in wells 1 & 2 (microscopy), zero signal in mock and untreated (3 & 4) | |||
* Harvest cells for flow cytometry | |||
Revision as of 15:00, 6 July 2013
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07/05/13
Lipo transfection
> Add 570 μL Opti-MEM to each 20 μL DNA sample Results (7/06/13)
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