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Magnesium Concentration

Magnesium is a required cofactor for thermostable DNA polymerases. Mg2+ in the PCR mixture stabilizes dsDNA and raises the Tm. Mg2+ concentration therefore is an important for controlling the specificity of the reaction. A low Mg2+ concentration requires more stringent base pairing in the annealing step. Too few Mg2+ ions result in a low yield of PCR product; too many Mg2+ ions increase the yield of non-specific products and promote misincorporation.
Real-Time PCR in Food Science
Edited by: David Rodríguez-Lázaro
An indispensable manual on real-time PCR for scientists in the food industry and for anyone involved in the detection of foodborne pathogens.
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Quantitative Real-time PCR in Applied Microbiology
Edited by: Martin Filion
Aimed specifically at microbiologists, this volume describes and explains the most important aspects of current real-time quantitative PCR (qPCR) strategies, instrumentation and software.
"useful book ... filled with valuable information" (Doodys); "an outstanding book" (Fungal Diversity) read more ...

Insufficient Mg2+ concentration in a PCR mixture can causes failure of the reaction. Excess magnesium (or the presence of manganese) will cause the fidelity of DNA polymerases to be reduced and may cause the generation of unwanted products. On a gel this can appear as a ladder or smear. The MgCl2 concentration should normally be between 1mM and 4mM. Since dNTPs sequester Mg2+ ions, a major change in the dNTP concentration in a rection would require a change in the concentration of MgCl2. Similarly, changing the KCl-based buffer concentration or any other component of the PCR mix may require adjustment of the Mg2+ concentration in the reaction mixture.

from PCR Troubleshooting: The Essential Guide see also PCR Troubleshooting and Optimization: The Essential Guide