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. 2014 Sep 4;33(21):2507–2520. doi: 10.15252/embj.201488398

Figure 3. Epigenetic instability of Bu-1a is dependent on the +3.5 G4 motif.

Figure 3

  1. The cartoons show the manipulation of the BU-1A locus in which the +3.5 G4 motif has already been deleted from both the BU-1A and BU-1B allele. Only the first three exons are shown along with the sequence reintroduced in the position of the original +3.5 G4 motif. (i) BU1A with the +3.5 kb G4 DNA deleted. (ii) BU1A with the +3.5 kb G4 DNA reintroduced. (iii) BU1A with the +3.5 kb G4 DNA reintroduced with two point mutations that abolish intramolecular G quadruplex formation in vitro (Fig 3B). (iv) BU1A with the +3.5 kb G4 DNA reintroduced inverted such that it would be present on the lagging strand template for a replication fork entering from the right of the locus. To the right is the fluctuation analysis of Bu-1a loss for each mutant. Each point represents the percentage of Bu-1alow variants in clones expanded for 20 generations. Red bar = median loss; whiskers = interquartile range; P-values that each distribution is different from control (i) calculated with Fisher's exact test with a bin size of 20%. NS = not significant (P < 0.001).
  2. Circular dichroism spectroscopy of the native BU-1 +3.5 G4 motif (red line) and a version mutated to prevent intramolecular G4 formation (blue line) carried out in KCl (solid line) and LiCl (dashed line).