Infecting Dinn-1 Cells with P4 DiNV for RNA
Infected Dinn-1 cells needed for time course for RNASeq, flies for day 0, 1, 3, and 5. Cells had already been plated a few days before here.
I used the 250ul aliquot of P4 DiNV, this was not diluted (see here), Rob and I decided that using 20ul seemed like a good amount for each plate of cells to have the max ability to see all cells infected.
- Thawed virus tube 81 on ice
- Got LN2 for day 0 cells
- Scraped up the wells for the day 0 cells with pipette tips inside the hood (Plate A)
- Transferred the solution to 1.5mL tubes per well
- Centrifuged tubes 1 min at 1000g to pellet the cells
- Removed the supernatant to separate tubes
- Flash froze the cell pellets:
| sample number | treatment | day |
|---|---|---|
| 1 | NA | 0 |
| 2 | NA | 0 |
| 3 | NA | 0 |
| 4 | NA | 0 |
For innoculating plates:
- Pipette mixed virus solution
- Added 20ul of P4 virus solution to wells C2, C3, C4, and C5 of plates B, C, and D
- Added 20ul of cell culture medium to wells B2, B3, B4, and B5 of plates B, C, and D
- Rocked plates back and forth a bit
- Placed plates back in the incubator
Freezing plates later on:
- On the day specified I followed the same process as above to freeze the cells
- Scraped up the wells for the day cells with pipette tips inside the hood
- Transferred the solution to 1.5mL tubes per well
- Centrifuged tubes 1 min at 1000g to pellet the cells
- Removed the supernatant to separate tubes
- Flash froze the cell pellets
All samples:
| sample number | treatment | day |
|---|---|---|
| 1 | NA | 0 |
| 2 | NA | 0 |
| 3 | NA | 0 |
| 4 | NA | 0 |
| 9 | CCM | 1 |
| 10 | CCM | 1 |
| 11 | CCM | 1 |
| 12 | CCM | 1 |
| 13 | 20ul P4 DiNV | 1 |
| 14 | 20ul P4 DiNV | 1 |
| 15 | 20ul P4 DiNV | 1 |
| 16 | 20ul P4 DiNV | 1 |
| 25 | CCM | 3 |
| 26 | CCM | 3 |
| 27 | CCM | 3 |
| 28 | CCM | 3 |
| 29 | 20ul P4 DiNV | 3 |
| 30 | 20ul P4 DiNV | 3 |
| 31 | 20ul P4 DiNV | 3 |
| 32 | 20ul P4 DiNV | 3 |
| 41 | CCM | 5 |
| 42 | CCM | 5 |
| 43 | CCM | 5 |
| 44 | CCM | 5 |
| 45 | 20ul P4 DiNV | 5 |
| 46 | 20ul P4 DiNV | 5 |
| 47 | 20ul P4 DiNV | 5 |
| 48 | 20ul P4 DiNV | 5 |
Sample info here