+ Site Statistics
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on LinkedInFollow on LinkedIn

+ Translate

Energy transfer cassettes for facile labeling of sequencing and PCR primers

Energy transfer cassettes for facile labeling of sequencing and PCR primers

Analytical Biochemistry 292(2): 188-197

Fluorescence energy transfer (ET) primers and terminators are the reagents of choice for multiplex DNA sequencing and analysis. We present here the design, synthesis and evaluation of a four-color set of ET cassettes, fluorescent labeling reagents that can be quantitatively coupled to a thiol-activated target through a disulfide exchange reaction. The ET cassette consists of a sugar-phosphate spacer with a FAM donor at the 3'-end, an acceptor linked to a modified T-base at the 5'-end of the spacer and a mixed disulfide for coupling to a thiol at the 5'-end. The acceptor dye emission intensities of ET labeled primers produced in this manner are comparable to commercial ET primers. The utility of our ET cassette-labeled primers is demonstrated by performing four-color capillary electrophoresis sequencing with the M13(-21)forward primer and by generating and analyzing a set of single-nucleotide-polymorphism-specific PCR amplicons.

(PDF emailed within 0-6 h: $19.90)

Accession: 010588028

Download citation: RISBibTeXText

PMID: 11355850

DOI: 10.1006/abio.2001.5069

Related references

Cassette labeling for facile construction of energy transfer fluorescent primers. Nucleic Acids Research 24(6): 1144-1148, 1996

Energy transfer primers: a new fluorescence labeling paradigm for DNA sequencing and analysis. Nature Medicine 2(2): 246-249, 1996

A three-wavelength labeling approach for DNA sequencing using energy transfer primers and capillary electrophoresis. Electrophoresis 19(8-9): 1403-1414, 1998

Synthesis of symmetrical multichromophoric Bodipy dyes and their facile transformation into energy transfer cassettes. Chemistry 16(21): 6346-6351, 2010

Fluorescent, through-bond energy transfer cassettes for labeling multiple biological molecules in one experiment. Journal of the American Chemical Society 125(48): 14668-9, 2003

Fluorescence energy transfer dye-labeled primers for DNA sequencing and analysis. Proceedings of the National Academy of Sciences of the United States of America 92(10): 4347-4351, 1995

Primers labeled with energy transfer coupled dyes for DNA sequencing. Official Gazette of the United States Patent & Trademark Office Patents 1206(2): 1340-1341, Jan 13, 1998

DNA sequencing with solid-phase-capturable dideoxynucleotides and energy transfer primers. Analytical Biochemistry 309(1): 35-39, October 1, 2002

Design and synthesis of fluorescence energy transfer dye-labeled primers and their application for DNA sequencing and analysis. Analytical Biochemistry. 231(1): 131-140, 1995

Correlations of structure and rates of energy transfer for through-bond energy-transfer cassettes. Journal of Physical Chemistry. A 110(1): 20-27, 2006

Ligation mediated fluorescent labeling of DNA sequencing primers. Nucleic Acids Research 25(4): 922-923, 1997

Enzymatic fluorescence and biotin labeling of primers for PCR sequencing. Methods in Molecular Biology 65: 23-28, 1996