Picked KAH79 and KAH126
Transfected each well of two 12-well plates with 1ug of plasmid DNA.
The cells looked way better than last time. Luc14s were about 80-90% confluent while the gal4-eeds were about 90-95% confluent. The media for the gal4s was starting to turn orange so I added 1mL of media to each well (added dox to the gal4s)
Step 0: Prep DNA
definitely g023, g034
for the third: in order of importance: g048, g046, g044, g025
need 3 tubes with 1ug/10ul each.
- 1.8ul KAH126 + 8.2ul elution solution
- 10ul elution sol'n
Step 1: Make mastermixes
MM1: Mix PLUS reagent with optimem
| PLUS reagent||1||25
MM 2: Lipo LTX and optimem
| Lipofectamine LTX||3||75
Step 2: Mix MM1 with DNA
- Add 91ul of MM1 to 1ug DNA in 10ul of Elution solution
- Incubate for 5 minutes at RT
Step 3: Mix in Lipo LTX
- Add 100ul of MM2 to DNA-MM1
- Incubate for 30 minutes at RT
Steo 4: Add complexes dropwise to each well
Everything went well, finished about 4:30.