Description:

IsoAmpIII 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® III Universal tHDA kit is based on a third-generation tHDA formula for more robust and specific amplification with reduced non-specific products due to primer-dimer. In some cases, rapid amplification may be achieved by adding more IsoAmp® III Enzyme Mix into the reaction (unpublished observation). The volume of IsoAmp® III Enzyme Mix has been reduced to 2 μL per reaction, which allows the user to add more enzymes (2 X or 3 X) for possible high-speed amplification. IsoAmp® III Enzyme Mix (Cat# H1020S) can be purchased separately.

IsoAmp® III Universal tHDA kit supports all reactions which are supported by IsoAmp® II Universal tHDA kit. Selected examples are included in this manual, including end-point tHDA and real-time quantitative HDA (qHDA) in both one-step and two-step formats. A Control Template and a set of specific primers are supplied for the positive control reaction.

Figure 2.Positive control assays with varying amount of Control Template.
Positive control assays were conducted following the one-step qHDA protocol. Control Template was used at 5 ng - 500 fg per assay in triplicate.

* Negative control reactions may produce primer-dimer, which can be distingushed from specific amplicons by gel-analysis or by melting-curve analysis.

Applications:

Used to amplify and detect short DNA sequences (80 - 120 bp)

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Advantages:
Use of helicase to separate DNA eliminates need for thermocycler
Reactions can be performed at constant temperature.
Works with a variety of templates, including bacterial genomic DNA, viral DNA, plasmid DNA and cDNA.
As low as a single copy of target DNA can be amplified under optimized conditions.
Can be used for real-time quantitative or/and reverse transcription HDA.

Kit Components:

Storage Conditions:

Quality Control:

Quality Assurance Statement:
Kit components were individually tested for optimal performance in tHDA.

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