PCR Guru by Najafov Ayaz; Hoxhaj Gerta; & Gerta Hoxhaj

Author:Najafov, Ayaz; Hoxhaj, Gerta; & Gerta Hoxhaj
Language: eng
Format: epub
Publisher: Elsevier Science & Technology

Figure 4.1 Annealing temperature gradient.

Decreasing the annealing temperature too low leads to nonspecific amplification (lane 1) and raising it too high leads to lack of any amplification.

Even though the main role of Mg++ ions is to serve as a cofactor for DNA polymerase, Mg++ ions bind to deoxynucleotidetriphosphates (dNTPs), primers, and template DNA too. A fraction of Mg++ ions that is added to the reaction will always be chelated by these PCR components. Therefore, whenever the concentration of any of these components is altered, in the case if the PCR result is not satisfactory, the first thing that has to be optimized is the Mg++.

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