 |
|
 |
|
|
| Home >
Products >
Polymerases >
Amplification and Cloning Technologies >
IsoAmp® tHDA Kit |
 | | | | | | Developed and produced by BioHelix Corp., a NEB-affiliated company. | |
| | |
 |
|
This product is currently
not available for online ordering.
|
This product has been discontinued and replaced by H0110
- Easy-to-use for assay development
- Use of helicase to separate DNA eliminates need for thermocycler
- Reactions can be performed at constant temperature
Description:
The IsoAmp® tHDA kit can be used to amplify and detect a short DNA sequence (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. As few as 10 copies of target DNA can be amplified by tHDA and detected by agarose gel electrophoresis, when optimized primers and buffer are used.
The IsoAmp tHDA kit is based on the thermophilic Helicase-Dependent Amplification (tHDA) method. 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. Thus DNA can be amplified at a single temperature without the need for thermocycling.
The IsoAmp tHDA kit provides the necessary reagents in a convenient 2X master mix format, including a thermostable DNA helicase, a DNA polymerase, dNTPs and other components. 10X Annealing Buffer is also provided for the initial hybridization of template and primers. Separate tubes of NaCl and MgSO4 are provided for further optimization of the amplification reaction. Control template and a set of amplification primers are supplied for a positive control reaction.
Figure 1: HDA technology.
Figure 2. Positive control assay of a 99 bp fragment with varying amount of Control Template. See manual for reaction conditions.
Applications:- Used to amplify and detect short DNA sequences (70 - 120 bp)
Advantages:- Supplied as a convenient 2X Master Mix format
- Works with a variety of templates, including bacterial genomic DNA, viral DNA, plasmid DNA and cDNA
- As low as 10 copies of target DNA can be amplified under optimized conditions
Kit Components:
Annealing Buffer (10X)
Control Forward Primer (5 μM)
Control Reverse Primer (5 μM)
Control Template (Plasmid: 1 ng/µl)
MgSO4 (100 mM)
NaCl (500 mM)
tHDA Mix (2X)
Storage Conditions
Storage Temperature:
-20°C
Notes
General notes:- Recommended storage conditions for the tHDA kit are -20°C for short-term (less than 6 months) storage and -80°C for storage greater than 6 months. This kit has not been optimized for amplification of human genomic DNA.
FAQs
- Can an alternate buffer system be used with the IsoAmp tHDA Kit?
- Can HDA with the IsoAmp tHDA Kit be performed at temperatures other than 64°C?
- Can I perform HDA with the IsoAmp tHDA Kit in a thermocycler?
- Can PCR primers be used with the IsoAmp tHDA Kit?
- Can the HDA reaction using IsoAmp tHDA Kit be run in smaller reaction volumes (less than 50μl)?
- Can the IsoAmp tHDA Kit amplify DNA sequences in excess of 120 bp?
- Can the IsoAmp tHDA Kit be used with Real-Time PCR machines?
- Does HDA with the IsoAmp tHDA Kit require heat denaturation?
- How much 100 mM MgSO4 should be added to the HDA reaction using the IsoAmp tHDA Kit?
- What is the detection limit for the IsoAmp tHDA Kit?
- What templates can the IsoAmp tHDA Kit be used with?
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Exonuclease Activity:
Incubation of 25 ml IsoAmp 2X tHDA Mix for 4 hours at 37°C in 50 µl reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 µg of a mixture of single and double-stranded [3H] E. coli DNA (200,000 cpm/µg) released < 0.05% of the total radioactivity.
Endonuclease Assay:
Incubation of 25 ml IsoAmp 2X tHDA Mix for 4 hours at 37°C in 50 µl reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 µg ΦX174 RFI DNA gave < 5% conversion to RF II.
Nuclease Activity:
Incubation of 25 ml IsoAmp 2X tHDA Mix for 16 hours at 37°C in 50 µl of reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 µg l DNA yielded a clear and sharp band on an agarose gel.
Legal
Licenses/Patents/Disclaimers:
NOTICE TO PURCHASER: This product is covered by pending U.S. and foreign patents. The purchase price of this product includes a limited, non-transferable license for research use only under the pending patents owned by New England Biolabs and BioHelix. No other license under these patents is conveyed expressly or by implication to the purchaser by the purchase of the product. The Product is not to be used for diagnostic purposes nor is it intended for human use. This product may not be resold, modified for resale, or used to manufacture commercial products without written approval of BioHelix Corp. Further information regarding this product may be obtained by contacting BioHelix at information@biohelix.com.
|
|
|
 |
|
|
