Gene mapping and gene enrichment by the avidin-biotin interaction: use of cytochrome-c as a polyamine bridge.
AUTOR(ES)
Sodja, A
RESUMO
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 diamine to the oxidized 2', 3' terminus of an RNA by Schiff base formation and BH-4 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.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=341990Documentos Relacionados
- Three-dimensional structures of avidin and the avidin-biotin complex.
- Avidin-biotin interactions at vesicle surfaces: adsorption and binding, cross-bridge formation, and lateral interactions.
- Molecular dynamics study of unbinding of the avidin-biotin complex.
- Application of the avidin-biotin method of gene enrichment to the isolation of long double-stranded DNA containing specific gene sequences.
- Rapid enrichment of HeLa transcription factors IIIB and IIIC by using affinity chromatography based on avidin-biotin interactions.
