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The FTA® cards were dried, then directly added to a polymerase chain reaction (PCR) mix and processed by PCR.
Oligonucleotides were removed from the PCR mix by PCR 96 cleanup plate (Millipore).
PCR mix and conditions were similar as described above, with the exception of only using 8 cycles for amplification.
PCR mix and conditions were similar as described above, with an exception of using 8 cycles for amplification.
BRCT regions were amplified by KOD Polymerase PCR (Supplementary Table S11) using Matchmaker Insert Check PCR Mix 2 (Clontech) for mutation identification by Sanger sequencing.
The incubated PCR mix was directly loaded onto the gel with the ladder (GeneDirex DM003-R500, 7 μl 100 3,000 bp).
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To improve amplification, Q-Solution (Qiagen, Hilden, Germany) was added to the RT-PCR mix.
Negative control was done as described with a PCR-mix lacking cDNA.
Additionally, the PCR-Mix contained 10 n M of the TaqMan-Probe.
Immunoprecipitated DNA was amplified by quantitative real-time PCR (ABI Power SYBR Green PCR mix).
For ABCD1 fragment amplification, long-range PCR was performed using the QIAGEN Long-range PCR mix.
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