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

Broker, T. R., Angerer, L. M., Yen, P. H., Hershey, N. D., Davidson, N. (1978) Electron microscopic visualization of tRNA genes with ferritin-avidin: biotin labels. Nucleic Acids Res, 5 (2). pp. 363-84. ISSN 0305-1048 (Print)0305-1048 (Linking)

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URL: http://www.ncbi.nlm.nih.gov/pubmed/345243
DOI: 10.1093/nar/5.2.363

Abstract

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 complementary DNA sequences present in a single stranded non-homology 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 microscopy specifically and accurately reveals the position of the tRNA gene, with a frequency of labeling of approximately 50%.

Item Type: Paper
Uncontrolled Keywords: 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
Subjects: Investigative techniques and equipment > microscopy > electron microscopy
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > tRNA
CSHL Authors:
Communities: CSHL labs
Depositing User: Matt Covey
Date Deposited: 12 Dec 2016 20:57
Last Modified: 08 Nov 2017 17:15
PMCID: PMC341989
Related URLs:
URI: http://repository.cshl.edu/id/eprint/33312

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