Extended Data Fig. 4: Transgenic H2B expression does not exacerbate CIN. | Nature Genetics

Extended Data Fig. 4: Transgenic H2B expression does not exacerbate CIN.

From: Reconstructing single-cell karyotype alterations in colorectal cancer identifies punctuated and gradual diversification patterns

Extended Data Fig. 4

a, The HIST1H2BC locus, coding for most abundant Histone H2B, was targeted to fluorescently tag H2B1C with Dendra2. Cas9 nickase (D10A) was targeted to the endogenous STOP codon, as well as both sites of the homology arms flanking the donor template. b, 18 days post electroporation, H2B-Dendra2 positive cells were isolated using FACS. Overall knock-in efficiency of 1.7% (not corrected for electroporation efficiency of ~10%). c, Similar CIN phenotypes of PDTO-9 expressing transgenic H2B-Dendra2 or carrying a HIST1H2BC-Dendra2 knock-in. 649 divisions across 34 organoids and 183 divisions across 27 organoids were scored from the transgenic and knock-in line respectively. Difference in scored chromatin error rates is statistically significant (Chi square test, p = 0.031), although the error rate in the knock-in line is likely underreported due to the lower brightness of endogenous H2B-Dendra2 expression. d, Fluorescent signal of H2B-Dendra was compared between PDTO-9 lines expressing transgenic H2B-Dendra2 (n = 95 cells, 19 organoids) or carrying a HIST1H2BC-Dendra2 knock-in (n = 100 cells, 20 organoids). Data are represented as box-and-whisker plots; boxes represent quartiles 2 and 3, the horizontal line represents the mean and whiskers represent minima and maxima within the 1.5× interquartile range.

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