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== Repeat Expansion PCR Test == * I repeat amplification by increasing amount of oligos to 5X, 10X, and 20X, and used the 100nM oligos as the template. I am not sure if the high concentration of oligos containing the majority of truncated oligos will interfere the PCR that much or not * I also increase extension time to 45 sec even 30 sec is enough to synthesize 200nt DNA fragment, and increase the second step PCR to 15 cycles (total = 20 cycles). This is just to monitor the qPCR curve of each concentration. ** 1. NTC ** 2. +ve control 1nM LC Sciences oligos ** 3. 5nM 12k CustomArray oligos (5X, added 1.25ul of 100nM in 25ul reaction) ** 4. 10nM 12k CustomArray oligos (10X, added 2.5ul of 100nM in 25ul reaction) ** 5. 20nM 12k CustomArray oligos (20X, added 5ul of 100nM in 25ul reaction) {| {{table}} | align="center" style="background:#f0f0f0;"|'''Components''' | align="center" style="background:#f0f0f0;"|'''1 rxn''' | align="center" style="background:#f0f0f0;"|'''5.5 rxn mix''' |- | 20nM seed oligoes||1.25||0.00 |- | eMIP_CA1_F (100uM)||0.10||0.55 |- | eMIP_CA1_R (100uM)||0.10||0.55 |- | 2x Kapa SYBG MM||12.50||68.75 |- | H2O||11.05||60.78 |- | Total||25.00||137.50 |} * Aliquot 23.75 '''Program'''<br> 95C 30sec -> (95C 5sec -> 52C 60sec-> 72C 45sec) x 5 -> (95C 5sec -> 60C 30sec-> 72C 45sec) x 15 -> 72C 2min -> 15C hold<br>
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