The isolation of specific chicken macrochromosomes by zonal centrifugation and flow sorting

Abstract

The nine largest macrochromosomes of the chicken, comprising about 80% of the genome, have each been isolated in yields of about 2 × 10⁶ chromosomes and approximately 80% purity, except for chromosome 4, which is only about 55% pure because it is heavily contaminated with sex chromosome Z. The chromosomes were first separated roughly according to size by zonal centrifugation on a sucrose gradient. Aliquots of the gradient fractions were then stained with ethidium bromide and analyzed by a flow cytometer-sorter. Electrostatic drop deflection was used to sort the desired chromosomes from chosen zonal gradient fractions. Each fraction was reexamined by flow analysis and by fluorescence microscopy. The DNA from these purified chromosomes has been used for gene mapping by molecular probe analysis.