Hybridization assays using as a label an expressible DNA fragment encoding firefly luciferase.
- UNCG Author/Contributor (non-UNCG co-authors, if there are any, appear on document)
- Norman H. Chiu, Assistant Professor (Creator)
- Institution
- The University of North Carolina at Greensboro (UNCG )
- Web Site: http://library.uncg.edu/
Abstract: We report the use of a new label, an expressible enzymecoding
DNA fragment, for nucleic acid hybridization
assays. The DNA label contains a firefly luciferase coding
sequence downstream from a T7 RNA polymerase promoter.
The target DNA (200 bp) is denatured and
hybridized simultaneously with two oligonucleotide probes.
One of the probes is immobilized in microtiter wells, via
the digoxigenin/anti-digoxigenin interaction, and the other
probe is biotinylated. After completion of the hybridization,
the hybrids are reacted with a streptavidin-luciferase
DNA complex. Subsequently, the solid-phase
bound DNA is expressed by coupled transcription/
translation. The synthesized luciferase catalyzes the
luminescent reaction of luciferin with O2 and ATP. The
luminescence is linearly related to the amount of target
DNA in the range of 5-5000 amol. The CVs obtained
for 20 and 100 amol of target are 6.5% and 10.8%,
respectively (n ) 4).
Hybridization assays using as a label an expressible DNA fragment encoding firefly luciferase.
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Created on 12/11/2012
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Additional Information
- Publication
- Language: English
- Date: 1996
- Keywords
- chemistry, biochemistry, DNA, enzyme-coding, DNA label, hybridization, firefly luciferase