Electron microscopic visualization of tRNA genes with ferritin-avidin: biotin labels

A method is described for indirect electron microscopic visualization and mapping of tRNA and other short transcripts hybridized to DNA. This method depends upon the attachment of the electron-dense protein ferritin to the RNA, the binding being mediated by the remarkably strong association of the e...

Full description

Saved in:
Bibliographic Details
Published in:Nucleic acids research 1978-02, Vol.5 (2), p.363-384
Main Authors: Broker, Thomas R., Angerer, Lynne M., Yen, Pauline H., Hershey, N. Davis, Davidson, Norman
Format: Article
Language:English
Subjects:
Avidin
Biotin
Chemical Phenomena
Chemistry
Chromosome Mapping
Coliphages
DNA, Bacterial - genetics
Escherichia coli - genetics
Ferritins
Genes
Microscopy, Electron - methods
RNA, Bacterial - genetics
RNA, Transfer - genetics
Citations: Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:A method is described for indirect electron microscopic visualization and mapping of tRNA and other short transcripts hybridized to DNA. This method depends upon the attachment of the electron-dense protein ferritin to the RNA, the binding being mediated by the remarkably strong association of the egg white protein avidin with biotin. Biotin is covalently attached to the 3′ end of tRNA using an NH2 (CH2) 5NH2 bridge. The tRNA-biotin adduct is hybridized to complementcrry DNA sequences present in a single stranded nonhomology loop of a DNA:DNA heteroduplex. Avidin, covalently crosslinked to ferritin is mixed with the heteroduplex and becomes bound to the hybridized tRNA-biotin. Observation of the DNA:RNA-biotin:avidin-ferritin complex by electron microsdopy specifically and accurately reveals the position of the tRNA gene, with a frequency of labeling of approximately 50%.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/5.2.363