Selection and Nuclear DNA Analysis of Cell Hybrids between Daucus carota and Oryza sativa

Abstract

In order to study the fate of the parental genomes in somatic cell hybrids between distantly related species, protoplasts from cultured cells of Daucus carota and Oryza sativa were fused. Selective conditions resulted, exclusively, in the growth of hybrid colonies which combined the capacity to multiply of carrot cells with the natural resistance to A2CA of rice cells. A methodology for measuring the relative contribution of the parental cells to the hybrid nuclear genome has been worked out. This is based both on hybridization of nuclear DNA bound to nitrocellulose filters (dot hybridization) with radioactively labelled nuclear DNA from one of the parents and on agarose gel fractionation of nuclear DNA digested with restriction endonucleases. The dot hybridization analysis, performed on one of the D. carota x O. sativa cell hybrids, showed that the major portion of the nuclear genome is homologous with the carrot partner, with rice contributing only a minor fraction, along with the selected resistance gene(s). The homology was confirmed after agarose gel fractionation of restriction endonuclease BamHI-digested nuclear DNA. Furthermore, strong homology at the level of gene expression between hybrid and carrot cells was shown by polyacrilamide gel electrophoresis of total soluble proteins.