Navigation Links
Preventing Carry-over Contamination with Uracil-DNA Glycosylase

The polymerase chain reaction (PCR) can amplify a single molecule over a billionfold. Thus, even minuscule amounts of a contaminant can be amplified and lead to a false positive result. Such contaminants are often products from previous PCR amplifications (carry-over contamination). Therefore, researchers have developed methods to avoid such contamination.

One common strategy is substituting dUTP for dTTP during PCR amplification, to produce uracil-containing DNA (U-DNA). Treating subsequent PCR reaction mixtures with Uracil-DNA Glycosylase (UNG) prior to PCR amplification and subsequent cleavage of apyriminic polynucleotides at elevated temperature (95C) under alkaline conditions (during the initial denaturation step) will remove contaminating U-DNA from the sample (see figure below). This method, of course, requires that all PCR-reactions in the lab have to be carried out with dUTP instead of dTTP.

Note the following when using dU-containing PCR products in downstream applications:
  • PCR products containing dU perform as well as those containing dT when used as hybridization targets or as templates for dideoxy sequencing.
  • PCR products containing dU can be cloned directly, if they are transformed into ungbacterial hosts.
  • A dU-containing substrate is readily digested by some common restriction enzymes (e.g. Eco RI and Bam HI), while others show reduced activity (e.g. Hpa I, Hind II, Hind III) on these substrates.
  • We do not recommend the use of dU
    '"/>


Source:


Page: All 1 2

Related biology technology :

1. Combine High Yield, Great Accuracy and the Prevention of Carry-over Contamination by Using the Novel Expand High FidelityPLUS PCR System
2. Mycoplasma Contamination Reduces the Effect of Lipid-Mediated Transfection of Mammalian Cells
3. Combat RNase Contamination in the Lab
4. The Fastest, Simplest, and Most Effective Way to Remove DNA Contamination
5. Testing Compressed Air Lines for Microbiological Contamination
6. Testing Compressed Air Lines for Microbiological Contamination
7. Uracil-DNA Glycosylase
8. Uracil-DNA Glycosylase, heat-labile
Post Your Comments:
(Date:9/23/2014)... 23, 2014 This is a ... of the Phenolic Resin industry in Global and ... industry, including definitions, applications and industry chain structure. ... are provided with a focus on history, developments, ... comparison between the international and Chinese situation is ...
(Date:9/23/2014)... de Reuil, France (PRWEB) September 23, 2014 ... to protection of laboratory personnel since 1968 and Burdinola, ... the development of a new filtered fume hood. ... above their laboratory fume hood, Burdinola will now offer ... for filtering a wide range of chemicals, best-in-class detection ...
(Date:9/23/2014)... Proove Biosciences , the commercial and research ... the association between specific genes and patients’ predisposition to ... Health Congress’ 27th Annual Scientific Program in Orlando, ... of ‘Embracing Change.’ Mental health, and healthcare as a ... challenges presented by the Affordable Care Act to the ...
(Date:9/23/2014)... Sept. 23, 2014 Research and ... "Proteomics Markets for Research and IVD Applications (Mass ... report to their offering. This ... applications, both in their research use and in ... the study of the structure and function of ...
Breaking Biology Technology:Phenolic Resin Industry (International, China) Development Policy and Plan Now Available at ReportsnReports.com 2Erlab Partners with Burdinola to Deliver an Innovative Filtered Fume Hood 2Proove Biosciences Will Be Exhibiting New Genetic Research at the 27th Annual US Psych Congress Scientific Program 2Proove Biosciences Will Be Exhibiting New Genetic Research at the 27th Annual US Psych Congress Scientific Program 3Proteomics Markets for Research and IVD Applications (Mass Spectrometry, Chromatography, Microarrays, Electrophoresis, Immunoassays, Other Technologies) 2Proteomics Markets for Research and IVD Applications (Mass Spectrometry, Chromatography, Microarrays, Electrophoresis, Immunoassays, Other Technologies) 3Proteomics Markets for Research and IVD Applications (Mass Spectrometry, Chromatography, Microarrays, Electrophoresis, Immunoassays, Other Technologies) 4
... The power to edit genes is as revolutionary, immediately ... Gutenberg,s printing press. And like Gutenberg,s invention, most DNA ... brilliant technology in its infancy. Now, Harvard researchers developing ... processing have rewritten the genome of living cells using ...
... Neogen Corporation (NASDAQ: NEOG ) announces ... , Tuesday, July 26, 2011 @ 11:00 ETWhere: , ... -- Simply log on to the web at the ... Corporation, +1-800-234-5333If you are unable to participate during the ...
... time that the hormone leptin acts in the brain to ... mystery. Now, new research published by Cell Press in the ... neurobiological mechanisms that may underlie the antiobesity effects of leptin. ... cells and acts at its receptor in the brain to ...
Cached Biology Technology:Editing the genome 2Editing the genome 3Modulation of inhibitory output is key function of antiobesity hormone 2
(Date:9/23/2014)... more carbon than the atmosphere and small changes ... effect on atmospheric greenhouse gas concentrations. A new ... concludes that climate warming does not accelerate ... storage, despite increases in ecosystem productivity. , ... Ecologist Dr. Christian Giardina, with the agency,s Institute ...
(Date:9/23/2014)... funds to support the next phase of its Tissue ... predicting drug safety and effectiveness. Researchers will collaborate over ... and combine them into an integrated system that can ... by the National Center for Advancing Translational Sciences (NCATS), ... in 2014 with additional support over the remaining two ...
(Date:9/23/2014)... tissues during the healing process, cells must be ... scientists believed this communication happened primarily through chemical ... the University of Pittsburgh have found that another ... not more crucial. The findings, published in this ... National Academy of Sciences , could lead to ...
Breaking Biology News(10 mins):Study helps assess impact of temperature on belowground soil decomposition 2NIH funds next phase of Tissue Chip for Drug Screening program 2NIH funds next phase of Tissue Chip for Drug Screening program 3NIH funds next phase of Tissue Chip for Drug Screening program 4The mechanics of tissue growth 2
... 20 years have suggested (though not unequivocally,confirmed) that semen ... sperm counts and,reduced sperm motility.(1) The explanations have been ... and to male smoking. Now, a new study,based on ... study cohorts, suggests that,exposure to several factors in utero ...
... with thinner roots can grow deeper, a trait which could ... New research, to be presented at the Society for Experimental ... have fewer cortical cells in the outer layer of their ... A research team headed by Prof Jonathan Lynch at the ...
... in Antarctica, is so dark, deep and cold that scientists ... place where nothing could live. However, work by Dr. ... sciences, and his colleagues has revealed a surprising variety of ... environments. A paper published June 26 in PLOS ONE ...
Cached Biology News:Early life and in utero factors found to influence testicular function in adulthood 2Early life and in utero factors found to influence testicular function in adulthood 3In subglacial lake, surprising life goes on 2In subglacial lake, surprising life goes on 3In subglacial lake, surprising life goes on 4In subglacial lake, surprising life goes on 5
Recombinant DNA polymerase from Thermus aquaticus expressed in E. coli; 5 units/L; 1 tube of 1, 000 units enzyme/tube w/Buffer II and MgCl2...
... - 5 units/µL ,• High specific activity ... Leaves an 'A' overhang ,• Processes ... very high activity in primer extension and ... quality of the enzyme, each lot has ...
Recombinant DNA polymerase from Thermus aquaticus expressed in E. coli; 5 units/uL; 25 tubes of 1, 000 units enzyme/tube w/ Buffer I...
... Gold® DNA Polymerase is a chemically modified ... chemical moiety is attached to the enzyme, ... the first ramp of thermal cycling (when ... temperatures), the enzyme is inactive. The result ...
Biology Products: