Figure 1.

DNA analysis. (a) Schematic representation of JAK2 gene region surrounding exon 14. The codon 617 mutation in JAK2V617F variant, together with the five SNPs defining the TCTT/GGCC subhaplotype of 46/1 haplotype are depicted, and their positions indicated by arrows. The polymorphic sites were found in linkage disequilibrium (see Text); hence, we focused on sequencing genomic PCR fragments encompassing the three sites surrounding the V617F mutation, and the mutation itself, using the two pairs of primers F2/R6 and F6/R7 (Supplementary Table 2), indicated at the bottom of the scheme. (b) Genetic characterization of the cell lines used. Genomic sequencing analysis led to confirm the JAK2 status regarding the amino acid at position 617, and to determine the three SNPs rs10974944, rs12343867 and rs1230895, as well as the deduced haplotypes. The predominant haplotype associated with the JAK2617V allele in SET-2 cells is indicated by an asterisk. Real-time PCR of a genomic region encompassing exon 8–intron 8 of JAK2, normalized to SDHA (succinate dehydrogenase complex subunit A) genomic fragment, served to determine JAK2 copy number in the tested cell lines, using the forward and reverse primers presented in Supplementary Table 2. DNA from peripheral blood lymphocytes of a healthy blood donor without any hematological anomaly was used as control.