10-20-22_AmpliconMM
- Gregg Randolph
Positive Control was not run, but primer dimers indicate rxns worked.
MasterMix (make for 40 plates in 50mL conical tube)
ul/rxn | Reagent | # of rxns | ul needed |
|---|---|---|---|
3 | 5X Kapa HiFi Buffer | 4100 | 12300 |
0.45 | 10M dNTPs | 4100 | 1845 |
0.3 | Kapa HiFi HotStart DNA Pol | 4100 | 1230 |
7.25 | HPLC H2O | 4100 | 29725 |
11 | Total Volume | 4100 | 45100 |
Aliquot 4,400 ul into 10 5ml tubes and label each #mm-dd-yy_AmpliconMM_#
Aliquot 11 ul into each well of an 8 pcr-tube strip. Add 4 ul IlluminaTest primers to 2. Add 4 ul 16STest primers to 2. Add 4 ul ITSTest primers to the next 2, Add 2 ul 16STest primers and 2 ul MC to the last 2..
Cap, Vortex, and spin down. Run on thermocycler program GSAF36:
Temp C | Cycles | Time |
|---|---|---|
95* | 1X | 3:00* |
98 | 36X | 0:30 |
62 | 36X | 0:30 |
72 | 36X | 0:30 |
72 | 1X | 5:00 |
4 | 1X | 0:00 |
No cleanup needed. Run resultant rxns on a D1000 High Sensitivity Tape.
Filename: 2022-10-21 - 09.59.03.HSD1000
All peaks are too small for proper products. They are most likely primer dimers.