Thermophilic Helicase-Dependent Amplification (tHDA) is a novel method for isothermal amplification of nucleic acids. Like PCR, the tHDA reaction selectively amplifies a target sequence defined by two primers. However, unlike PCR, tHDA uses an enzyme called a helicase to separate DNA, rather than heat. This allows DNA amplification without the need for thermocycling. The tHDA reaction can also be coupled with reverse transcription for RNA analysis.IsoAmp® II Universal tHDA kit can be used to amplify and detect short DNA sequences (70 bp - 120 bp) at a constant temperature. The kit can be used with a variety of templates, including microbial genomic DNA, viral DNA, plasmid DNA, and cDNA. A single copy of target DNA can be amplified by tHDA and detected by agarose gel electrophoresis when optimized primers and buffer are used. IsoAmp® II Universal tHDA kit is based on a second-generation thermophilic Helicase-Dependent Amplification platform. The reactions supported by IsoAmp® II Universal tHDA kit include tHDA, reverse transcription HDA (RT-HDA), real-time quantitative HDA (qHDA) and real-time quantitative RT-HDA (qRT-HDA), from a single reaction buffer. IsoAmp® II Universal tHDA kit contains an enzyme mix, dNTP solution and buffer, allowing flexibility in reaction setup. A Control Template and a set of specific primers are supplied for the positive control reaction. Recommended storage conditions for the tHDA kit are -20°C for storage shorter than 6 months and -80°C for storage greater than 6 months. Please avoid repeated freeze-thaw cycles as this may decrease performance of the kit.Figure 1.HDA Technology.Figure 2.Positive control assays with varying amount of Control Template. A, two-step tHDA positive control with 10 pg - 1 ng Control Template. B, two-step qHDA positive control using 500 fg - 5 ng, in triplicate.
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