Heterochromatin study demonstrating the non‐linearity of fluorometry useful for calculating genomic base composition

• Published in: Cytometry (New York, N.Y.) Vol. 14; no. 6; pp. 618 - 626
• Main Authors: Godelle, Bernard, Cartier, Delphine, Marie, Dominique, Brown, Spencer C., Siljak‐Yakovlev, Sonja
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.08.1993; Wiley-Liss
• Subjects: Adenine - analysis; Analytical, structural and metabolic biochemistry; AT; Base Composition; Benzimidazoles; Biological and medical sciences; bisbenzimide Hoechst; Cell Nucleus - chemistry; Crepis praemorsa complex; Cytosine - analysis; DNA - analysis; DNA - genetics; DNA amount; ethidium bromide; Flow Cytometry - methods; fluorochrome banding; Fluorometry - methods; Fundamental and applied biological sciences. Psychology; Genome; Guanine - analysis; Heterochromatin - chemistry; Karyotyping; Mathematics; mithramycin; Nucleic acids; Nucleic bases, nucleotides; Plants - genetics; Plicamycin; Thymine - analysis; Flow cytometry; Vegetals; Compositae; Heterochromatin; Nucleic base; Analysis method; Fluorescent labelling; Dicotyledones; DNA; Angiospermae; Spermatophyta; Genome; Nucleic acid; Quantitative analysis
• ISSN: 0196-4763; 1097-0320
• DOI: 10.1002/cyto.990140606

A novel procedure for calculating basepair frequencies in whole genomes is reported. This has been developed during a study of the role of heterochromatin in microevolution. Closely related species of the Crepis praemorsa complex have similar karyotypes but for their heterochromatin. The changes in relative AT frequency between species have been attributed to heterochromatin sequences by in situ banding of chromosomes with two base‐specific fluorochromes. The absolute genome size of species, measured by cytofluorometry, correlated positively with increased karyotypic heterochromatin, as did the proportion of AT bases in the DNA. However, the determination of base content has called for a curvilinear interpretation of data obtained with two base‐specific fluorochromes (bisbenzimide Hoechst 33342 and mithramycin), in contrast to the commonly assumed but erroneous direct relationship between fluorescence intensity and base content. Essentially, the fluorochromes' requirements for a sequence of certain basepairs lead to the notion of Coefficients or' Overspecificity: the result is a simple for mula for calculating the AT proportion in a genome relative to a reference species from cytometric data, taking account of ligand binding statistics. These statistics and probabilities of oligonucleotide binding are essentially the same. © 1993 Wiley‐Liss, Inc.