Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction
Abstract
Preparation of .sup.13 C/.sup.15 N-labeled DNA oligomers using the polymerase chain reaction (PCR). A PCR based method for uniform (.sup.13 C/.sup.15 N)-labeling of DNA duplexes is described. Multiple copies of a blunt-ended duplex are cloned into a plasmid, each copy containing the sequence of interest and restriction Hinc II sequences at both the 5' and 3' ends. PCR using bi-directional primers and uniformly .sup.13 C/.sup.15 N-labeled dNTP precursors generates labeled DNA duplexes containing multiple copies of the sequence of interest. Twenty-four cycles of PCR, followed by restriction and purification, gave the uniformly .sup.13 C/.sup.15 N-labeled duplex sequence with a 30% yield. Such labeled duplexes find significant applications in multinuclear magnetic resonance spectroscopy.
- Inventors:
-
- Los Alamos, NM
- Santa Fe, NM
- Issue Date:
- Research Org.:
- Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)
- OSTI Identifier:
- 873843
- Patent Number(s):
- 6258567
- Assignee:
- Regents of University of California (Los Alamos, NM)
- Patent Classifications (CPCs):
-
C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS
- DOE Contract Number:
- W-7405-ENG-36
- Resource Type:
- Patent
- Country of Publication:
- United States
- Language:
- English
- Subject:
- preparation; 13c; 15n-labeled; oligomers; polymerase; chain; reaction; 13; 15; n-labeled; dna; pcr; based; method; uniform; -labeling; duplexes; described; multiple; copies; blunt-ended; duplex; cloned; plasmid; copy; containing; sequence; restriction; hinc; ii; sequences; bi-directional; primers; uniformly; dntp; precursors; generates; labeled; twenty-four; cycles; followed; purification; 30; yield; significant; applications; multinuclear; magnetic; resonance; spectroscopy; resonance spectroscopy; labeled dna; chain reaction; magnetic resonance; polymerase chain; nuclear magnetic; based method; containing multiple; /435/999/
Citation Formats
Chen, Xian, Gupta, Goutam, and Bradbury, E Morton. Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction. United States: N. p., 2001.
Web.
Chen, Xian, Gupta, Goutam, & Bradbury, E Morton. Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction. United States.
Chen, Xian, Gupta, Goutam, and Bradbury, E Morton. Mon .
"Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction". United States. https://www.osti.gov/servlets/purl/873843.
@article{osti_873843,
title = {Preparation of 13C/15N-labeled oligomers using the polymerase chain reaction},
author = {Chen, Xian and Gupta, Goutam and Bradbury, E Morton},
abstractNote = {Preparation of .sup.13 C/.sup.15 N-labeled DNA oligomers using the polymerase chain reaction (PCR). A PCR based method for uniform (.sup.13 C/.sup.15 N)-labeling of DNA duplexes is described. Multiple copies of a blunt-ended duplex are cloned into a plasmid, each copy containing the sequence of interest and restriction Hinc II sequences at both the 5' and 3' ends. PCR using bi-directional primers and uniformly .sup.13 C/.sup.15 N-labeled dNTP precursors generates labeled DNA duplexes containing multiple copies of the sequence of interest. Twenty-four cycles of PCR, followed by restriction and purification, gave the uniformly .sup.13 C/.sup.15 N-labeled duplex sequence with a 30% yield. Such labeled duplexes find significant applications in multinuclear magnetic resonance spectroscopy.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Mon Jan 01 00:00:00 EST 2001},
month = {Mon Jan 01 00:00:00 EST 2001}
}
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