The distirubtion of active genes (globin) and inactive genes (keratin) in fractionated chicken erythroid chromatin

Abstract

Repeatable fractionation of sheared chromatin from purified populations of chicken erythroid cells has been achieved, based on the Bio-Gel procedure of Janowski et al. ((1972) Karolinska Symp. 5, 112). For reticulocytes, 3-5% of chromatin DNA is excluded from Bio-Gel A-50 m (peak I) and over 90% elutes in the included volume of the column(peak II). Peak I material has a higher protein/DNA ratio than peak II chromatin and the two fractions have melting profiles characteristic of "active" and "inactive" chromatin, respectively. In cells prelabeled with [3H]uridine or [3H]leucine there was very pronounced preferential association of radioactivity with the "active" peak I chromatin. The distribution of "active" (globin) and "inactive" (keratin) gene sequences in the DNA of fractions from peak I and peak II chromatin was determined with complimentary DNA (cDNA) probes to chicken globin mRNA and chicken feather keratin mRNA. While slight enrichment for globin gene sequences was found in peak I (relative to DNA in these fractions), some 80% of the total globin hybrid formed was found in peak II fractions. Experiments with the keratin cDNA probe showed that these genes were equally distributed in both chromatin fractions rather than being confined to the "inactive" peak II material. The hybridization data in particular question the validity of claims for fractionation of chromatin into "active" and "inactive" material.