Invitrogen platinum pcr supermix hi fi

Bi-directional Sanger sequencing was performed over the observed MED12 mutation hotspot (G44) on all tumor samples. Ten nanograms of genomic DNA was used as template in PCR amplifications with Invitrogen Platinum PCR Supermix Hi-Fi (Life Technologies, Foster City, CA) with the suggested initial denaturation and cycling conditions. Primer sequences were as previously reported (14 , 15 (link)) with the addition of universal M13 adaptors (M13 forward: TGTAAAACGACGGCCAGT and M13 reverse: CAGGAAACAGCTATGACC). PCR products were subjected to bidirectional Sanger sequencing for both primer pairs by the University of Michigan DNA Sequencing Core after treatment with ExoSAP-IT (GE Healthcare) and sequences were analyzed using SeqMan Pro software (DNASTAR, Madison, WI).

Bidirectional Sanger sequencing was performed over the prioritized mutations with variant allele frequencies >0.15 on all tumor samples. Genomic DNA (10ng) was used as template in PCR amplifications with Invitrogen Platinum PCR Supermix Hi-Fi (Life-Technologies) with the suggested initial denaturation and cycling conditions. PCR products were subjected to bidirectional Sanger sequencing for both primer pairs by the University of Michigan DNA Sequencing Core after treatment with ExoSAP-OT (GE Healthcare) and sequences were analyzed using SeqMan Pro Software (DNASTAR).