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Inactivation of Contaminating Templates in PCR Using High Intensity Ultraviolet Light


Introduction
Amplification of DNA through the polymerase chain reaction (PCR) is a highly sensitive method capable of copying millions of DNA molecules from just a single molecule.1 Because of the extreme sensitivity of this method, any carry-over product from previous reactions can result in false positives or extra bands. Carry-over contamination can be avoided by using positive displacement pipettes, sterile and autoclaved reagents, and dedicated areas for template preparation away from template amplification.

In addition, previous reports have noted that UV light is also an effective means of preventing carry-over contamination.2,3,4 High intensity UV light induces the formation of thymine dimers which block the progress of the Taq polymerase, and thus contaminating carry-over products are not amplified. In this bulletin, we describe a modified sterilization method using the GS Gene Linker UV chamber to destroy contaminating amplicon or carry-over molecules in microcentrifuge tubes prior to amplification.


Materials and Methods
Amplification of DNA was performed using genomic DNA from lambda phage and purified amplified human β-actin DNA. All of the tubes used in the amplification of a 500 bp fragment of lambda phage contained 50 ng of total lambda genomic DNA suspended in 5 l of distilled water. All of the tubes for the amplification of the amplicon DNA contained 20 ng of amplified β-actin product suspended in 2 l of distilled water. For the experimental UV tests, tubes were placed in the middle of the GS Gene Linker oven and were either capped and placed on their side or kept open and held in
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