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Fig. 1 | Acta Neuropathologica Communications

Fig. 1

From: H3F3A mutant allele specific imbalance in an aggressive subtype of diffuse midline glioma, H3 K27M-mutant

Fig. 1

Genetic analyses of H3F3A gene mutation using ddPCR and Sanger sequencing. a Results of ddPCR using various mixture DNA consisting of H3F3A K27M and wild-type H3F3A. Upper dot plots indicate wild-type H3F3A DNA dots (green) and lower dot plots indicate H3F3A K27M DNA dots (blue). The x-axis is the number of droplets including PCR products. The y-axis shows the intensity of HEX and FAM. The pink line shows the threshold (HEX: 1700, FAM: 2400) b Correlation of VAFs calculated by ddPCR with expected VAF. The x-axis is the expected VAF and the y-axis is VAF calculated by ddPCR (P < 0.01, r2 = 0.99). c 2D cluster plot of droplet fluorescence of H3F3A K27M and wild-type H3F3A dots in more than 50% VAF of H3F3A K27M case (case 12). FAM positive and HEX negative droplets (blue) include H3F3A K27M. HEX positive and FAM negative droplets (green) include wild-type H3F3A. VAF of H3F3A K27M was calculated as (blue dot numbers/blue dot numbers + green dot numbers). d Sanger sequencing of lower than 50% VAF case (left; 13.1% of VAF; case 8) or more than 50% VAF case (right; 79.9% of VAF; case 12). Hot spots of H3F3A K27M mutation are indicated by the red arrows

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