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Nucleic Acids Research, 1978, Vol. 5, No. 2 385-401
© 1978

Articles

Gene mapping and gene enrichment by the avidin-biotin interaction: use of cytochrome-c as a polyamine bridge

Ann Sodja and Norman Davidson

Division of Chemistry and Chemical Engineering, California Institute of Technology Pasadena, CA 91125, USA

Received October 31, 1977. A modification of previously described methods of electron microscopic gene mapping and of gene enrichment based on the avidin-biotin interaction is presented. The modification consists of coupling cytochrome-c instead of pentane tane diamine to the oxidized 2', 3' terminus of an RNA by Schiff base formation and Formula reduction. The RNA-cytochrome-c conjugate is purified by a simple chromatographic procedure; several biotins are attached to the cytochrome moiety by acylation. The extended arm between biotin and RNA gives efficient electron microscopic gene mapping of DNA:RNA-biotin hybrids with avidin-ferritin and avidin-polymethacylate sphere labels and efficient gene enrichment by buoyant banding of DNA:RNA-biotin:avidin-spheres in CsCl. A 1400 fold enrichment (thus, 25% pure) and a 90% yield of long Drosophila DNA strands with 5S RNA genes is achieved.


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