Non-Template Addition

Non-Template Addition

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image of split peaks on an electropherogram

If too much input DNA is added to an amplification reaction, the polymerase may be unable to complete the extension for all amplicons.03 Non-template addition results in a PCR product that is one base pair longer than the actual target sequence. When the polymerase is unable to complete the adenine addition on all products, this results in what is commonly referred to as split peaks (+A/-A peaks).01 To minimize split peaks, the extension phase of the PCR process is designed to drive the addition of adenine, ensuring that all amplicons are the same length. Kit manufacturers have also developed primer sequences that encourage adenine addition.

Click here to read more about PCR in Subject 01, Module 02.

Note:

Samples displaying -A can be diluted with buffer and re-extended in the thermal cycler.

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