A method for gene enrichment based on the avidin-biotin interaction. Application to the Drosophila ribosomal RNA genes.
A method of enriching, from the total DNA of an organism, for long DNA strands carrying a particular gene is described. The purified RNA corresponding to the gene is covalently attached to biotin via a cytochrome c bridge. This modified RNA is hybridized to the total DNA. Those DNA strands which hybridize are separated from all the other DNA, using the avidin-biotin interaction, by one of two methods. Avidin is covalently attached to submicroscopic polymer spheres; the complexes of avidin spheres with the DNA: RNA-biotin hybrids band in CsCl at a much lower buoyant density than does free DNA. Alternatively, the DNA:RNA-biotin hybrids are isolated by affinity chromatography on an avidin-solid support column. These methods have been used to prepare long single strands of Drosophila ribosomal DNA (rDNA) in high yield and 42 to 80% pure.
D000818:Animals; D001360:Avidin / Q000378:metabolism*; D001665:Binding Sites; D001710:Biotin / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D003574:Cytochrome c Group / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004331:Drosophila melanogaster / Q000378:metabolism*; D005796:Genes; D008854:Microscopy, Electron; D009690:Nucleic Acid Conformation; D009693:Nucleic Acid Hybridization; D009695:Nucleic Acid Renaturation; D010047:Ovalbumin / Q000031:analogs & derivatives*; D010957:Plasmids; D011485:Protein Binding; D012335:RNA, Ribosomal / Q000378:metabolism*; D014158:Transcription, Genetic
D003574:Cytochrome c Group; D012335:RNA, Ribosomal; D001360:Avidin; D001710:Biotin; D010047:Ovalbumin; D004247:DNA