| Cell Line Species/Tissue: |
Rat
/
Brain |
| Transfection Reagent: |
TransMessenger |
Nucleic Acid:
|
siRNA (dsRNA) |
Growth Medium:
|
DMEM / Neurobasal, B27 supplement + 0.5nM glutamine |
Percent Serum (%):
|
5 % |
Reporter System:
|
|
Plasmid Purification Method:
|
|
Plate Format:
|
6-well plate |
Number of Cells:
|
12.000-20.000 qcm poly L-lysin-coated coverslips |
Percent Confluence(%):
|
|
Amount of Nucleic Acid (µg):
|
1 µg |
Amount of Enhancer (µl):
|
2 µl |
Amount of Reagent (µl):
|
4 µl |
Complex Incubation on Cells (hrs):
|
2 h |
Analysis Performed Post-Transfection (hrs):
|
48-68 h |
Transfection Efficiency (%):
|
70-80% |
| Knockdown Efficiency (%): |
4 fold decrease in flourescent intensity |
| |
| Any modifications to the protocol?:
|
| |
| Notes: |
| The Transfection complex was diluted in 900 µl of Neurobasal and was added directly to the cells; it was replaced with Neurobasal after 2 h. The results were confirmed in three independent experiments. |
| References: |
| Krichevsky, A. M. and Kosik, K. S., PNAS (2002), vol. 99, no. 18, 11926–11929, RNAi functions in cultured mammalian neurons |