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. 2021 Jul 9;12:4202. doi: 10.1038/s41467-021-24451-8

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 6 — a Illustration of a series of haploid and diploid strains carrying a single additional copy of increasingly shorter fragments of promoters driving mCitrine expression, each truncated from the 5′-end. b mCitrine concentration at 60 fL normalized on maximum concentration of the respective promoter (upper panel) and VDP of mCitrine (bottom panel) determined by flow cytometry for the respective promoter truncations of the HTB1 promoter (dark blue circles) and the HHF1 promoter (light blue squares) driving mCitrine expression, integrated in haploid cells. Concentrations were estimated from a linear fit to the double logarithmic dependence of concentration on cell volume, VDPs were determined as the slope of the linear fit (fit through $n_{full}^{HTB 1 prom} = 12$ , $n_{750 bp}^{HTB 1 prom} = 15$ , $n_{600 bp}^{HTB 1 prom} = 15$ , $n_{450 bp}^{HTB 1 prom} = 15$ , $n_{300 bp}^{HTB 1 prom} = 15$ , and $n_{full}^{HHF 1 prom} = 12$ , $n_{600 bp}^{HHF 1 prom} = 15$ , $n_{450 bp}^{HHF 1 prom} = 15$ , $n_{300 bp}^{HHF 1 prom} = 15$ , $n_{150 bp}^{HHF 1 prom} = 15$ biological replicates). Error bars in the upper panel are derived by error propagation of the 95% confidence interval of the linear fit at 60 fL. In the bottom panel, error bars show the standard error of the VDPs. c VDP of mCitrine in haploid (blue filled circles) and diploid (green open squares) cells (upper panel) and mCitrine concentration at 60 fL in diploids compared to the concentration in haploids (bottom panel) determined by flow cytometry. Left shows results for the HTB1 promoter truncations, right shows results for the HHF1 promoter truncations. Concentrations were estimated from a linear fit to the double logarithmic dependence of concentration on cell volume, VDPs were determined as the slope of the linear fit (fit through $n_{full, haploid}^{HTB 1 prom} = 27$ , $n_{full, diploid}^{HTB 1 prom} = 18$ , $n_{450 bp, haploid}^{HTB 1 prom} = 27$ , $n_{450 bp, diploid}^{HTB 1 prom} = 27$ , $n_{300 bp, haploid}^{HTB 1 prom} = 27$ , $n_{300 bp, diploid}^{HTB 1 prom} = 27$ , and $n_{full, haploid}^{HHF 1 prom} = 27$ , $n_{full, diploid}^{HHF 1 prom} = 18$ , $n_{450 bp, haploid}^{HHF 1 prom} = 27$ , $n_{450 bp, diploid}^{HHF 1 prom} = 18$ , $n_{300 bp, haploid}^{HHF 1 prom} = 27$ , $n_{300 bp, diploid}^{HHF 1 prom} = 17$ biological replicates). Error bars in the upper panels show the standard error of the VDPs. In the bottom panel, error bars indicate the 2.5- and 97.5-percentiles around the median concentration ratio, determined from 10,000 bootstrap samples. d VDP of mCitrine in haploid (blue filled circles) and diploid (green open squares) cells (upper panel) and mCitrine mRNA concentration at 60 fL in diploids compared to the concentration in haploids (bottom panel) determined by RT-qPCR for HTB1 and HHF1 promoter truncations driving mCitrine expression. Concentrations were estimated from a linear fit to the double logarithmic dependence of concentration on cell volume, VDPs were determined as the slope of the linear fit (fit through $n_{450 bp, haploid}^{HTB 1 prom} = 15$ , $n_{450 bp, diploid}^{HTB 1 prom} = 14$ , $n_{300 bp, haploid}^{HTB 1 prom} = 16$ , $n_{300 bp, diploid}^{HTB 1 prom} = 12$ , and, $n_{450 bp, haploid}^{HHF 1 prom} = 12$ , $n_{450 bp, diploid}^{HHF 1 prom} = 12$ , $n_{300 bp, haploid}^{HHF 1 prom} = 11$ , $n_{300 bp, diploid}^{HHF 1 prom} = 12$ biological replicates). Error bars in the upper panel show the standard error. Error bars in the bottom panel indicate the 2.5- and 97.5-percentiles around the median concentration ratio, determined from 10,000 bootstrap samples.