Matt:LabNotes/2014-10-6
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Testing Agi26k0gap + suppressor Oligos & Agi26k20gap
Sample
- Agi26k0gap Control: PGP1F Fix 5.19.14 S2
- Agi26k0gap + suppressors: PGP1F Fix 5.19.14 S3
- Agi26k20gap: Fix 10.1.14
Procedure
- Agi26k0gap Control
Components | Volume |
5 U/ul Ampligase | 10 |
10X Ampligase Buffer | 10 |
Agi26k0gap | 35 |
H2O | 45 |
Total | 100 |
- Agi26k0gap + suppressors
- Combined 1ul of each 100uM 6 suppressor oligos in 1ml total to make 600nM oligo mix (100nM each suppressor oligo)
Components | Volume |
5 U/ul Ampligase | 10 |
10X Ampligase Buffer | 10 |
Agi26k0gap | 35 |
600 nM (100nM each) supp_Mix | 5 |
H2O | 40 |
Total | 100 |
- Agi26k20gap
Components | Volume |
5 U/ul Ampligase | 10 |
10X Ampligase Buffer | 10 |
Agi26k20gap | 77 |
25mM dNTP | 1 |
10 U/ul KlenTaq | 2 |
H2O | 0 |
Total | 100 |
- Incubate at 37C for 30 min
- Incubate at 60C for 24 hrs