Breeding was performed in the IGR or CINJ animal facilities. Tyr::CreERT2 and Ptenfl/fl mice were purchased from JAX (JAX012328 and JAX004597, respectively). BRafCA/CA mice were obtained from McMahon's laboratory.49 (link)
Atg7fl/fl mice were provided by Kamatsu's laboratory.50 (link)
By the age of 3 weeks, pup tails were partially cut for genotyping. DNA was extracted from using the DNAeasy Blood and Tissue Kit (Quiagen). DNA was then amplified by PCR in Micro-Amp 96-well reaction plate (Applied Biosystems), using REDExtract-N-Amp PCR ready Mix (Sigma). The primers used were 5′-GCG GTC TGG CAG TAA AAA CTA TC-3′, 5′-GTG AAA CAG CT TGC TGT CAC TT-3′, 5′-CTA GGC CAC AGA ATT GAA AGA TCT-3′ and 5′-GTA GGT GGA AAT TCT AGC ATC ATC C-3′ for Tyr-CRE, 5′-AAG CAC TCT GCG AAC TGA G-3′ and 5′-AAG TTT TTG AAG GCA AGA TGC-3′ for PTEN, 5′-TGA GTA TTT TTG TGG CAA CTG C-3′ and 5′-CTC TGC TGG GAA AGC GGC-3′ for BRAF, 5′-TGG CTG CTA CTT CTG CAA TGA TGT-3′ and 5′- CAG GAC AGA GAC CT CAG CTC CAC-3′ for ATG7 (JAX database).49,50 (link) PCR products were finally loaded on agarose gels and revealed, as previously described (JAX database).
Atg7fl/fl mice were provided by Kamatsu's laboratory.50 (link)
By the age of 3 weeks, pup tails were partially cut for genotyping. DNA was extracted from using the DNAeasy Blood and Tissue Kit (Quiagen). DNA was then amplified by PCR in Micro-Amp 96-well reaction plate (Applied Biosystems), using REDExtract-N-Amp PCR ready Mix (Sigma). The primers used were 5′-GCG GTC TGG CAG TAA AAA CTA TC-3′, 5′-GTG AAA CAG CT TGC TGT CAC TT-3′, 5′-CTA GGC CAC AGA ATT GAA AGA TCT-3′ and 5′-GTA GGT GGA AAT TCT AGC ATC ATC C-3′ for Tyr-CRE, 5′-AAG CAC TCT GCG AAC TGA G-3′ and 5′-AAG TTT TTG AAG GCA AGA TGC-3′ for PTEN, 5′-TGA GTA TTT TTG TGG CAA CTG C-3′ and 5′-CTC TGC TGG GAA AGC GGC-3′ for BRAF, 5′-TGG CTG CTA CTT CTG CAA TGA TGT-3′ and 5′- CAG GAC AGA GAC CT CAG CTC CAC-3′ for ATG7 (JAX database).49,50 (link) PCR products were finally loaded on agarose gels and revealed, as previously described (JAX database).
