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. 2014 Jan 7;289(7):3869–3875. doi: 10.1074/jbc.C113.532267

FIGURE 2.

FIGURE 2.

Exosomes contain mutated KRAS and p53 DNA. A, by PCR, we amplified a 466-bp fragment of KRAS spanning exon 2 and intron 2 and a 1564-bp fragment of p53 spanning 4 exons and 3 introns. B, Sanger sequencing of genomic DNA from Panc-1 cells and corresponding exosomes revealed the same heterozygous mutation of KRAS on codon 12 (GGT to GAT) and the similar homozygous mutation of p53 on codon 273 (CGT to CAT). T3M4 cells and corresponding exosomes displayed the same homozygous mutation of p53 on codon 220 (TAT to TGT). C, PCR amplification provided evidence for long fragments of DNA in circulating exosomes from two healthy donors and two patients with pancreatic cancer. We could retrieve a 466-bp fragment of KRAS DNA and 609-bp fragment of p53 DNA spanning exons 7 and 8 and intron 7. When serum samples depleted of exosomes were subjected to PCR, no KRAS or p53 amplicon could be detected. PDAC, pancreatic ductal adenocarcinoma. D, Sanger sequencing of serum exosome-derived DNA was able to detect DNA with a KRAS mutation on codon 22. In a second patient, Sanger sequencing revealed a KRAS mutation on codon 12 and a p53 mutation on codon 273.