Received gBlocks RibA, RibD, RibE, RibT25 and RibT48 and amplification oligos from IDT.
Dilution in water and PCR amplification, using the following protocol:
1 μL gBlock (0.1 to 1ng)
1 μL forward primer (10 μM)
1 μL reverse primer (10 μM)
22 μL DNAse/RNAse free water
25 μL LifeTech MasterMix (2X)
RibA + o15.001 (GCGCCCGAAGACTTATGCAG) + o15.002(GGCCCCGCGCATATGAAG)
RibD + o15.003(CGCTATAGAAGACTTGAGAAGATCTG) + o15.004(GCGCGGCACCACATATGAAG)
RibE + o15.005(CGGCTATAGAAGACTTGCGC) + o15.006(CGCGCCGGCATATGAAGA)
RibT25 + o15.007(CCGCGTATAGAAGACTGCTAGA) + o15.008(CAGCAGCATATGAAGACAACCC)
RibT48 + o15.009(GCGGTATAGAAGACTGCTAGAGA) + o15.010(CAGCAGCATATGAAGACAACCC)
For each PCR reaction, a negative control without matrix DNA was prepared.
30sec at 98°C
12 cycles
10sec at 98°C
30sec at 52°C
1min at 72°C
10min at 72°C
until opening at 12°C
Friday, Jul. 24
