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Home > Products > Polymerases > Amplification and Cloning Technologies > IsoAmp® tHDA Kit > FAQ IsoAmp® tHDA Kit FAQ See the Polymerases FAQ also. Q1: Can an alternate buffer system be used with the IsoAmp tHDA Kit? Q2: Can HDA with the IsoAmp tHDA Kit be performed at temperatures other than 64°C? Q3: Can I perform HDA with the IsoAmp tHDA Kit in a thermocycler? Q4: Can PCR primers be used with the IsoAmp tHDA Kit? Q5: Can the HDA reaction using IsoAmp tHDA Kit be run in smaller reaction volumes (less than 50μl)? Q6: Can the IsoAmp tHDA Kit amplify DNA sequences in excess of 120 bp? Q7: Can the IsoAmp tHDA Kit be used with Real-Time PCR machines? Q8: Does HDA with the IsoAmp tHDA Kit require heat denaturation? Q9: How much 100 mM MgSO4 should be added to the HDA reaction using the IsoAmp tHDA Kit? Q10: What is the detection limit for the IsoAmp tHDA Kit? Q11: What templates can the IsoAmp tHDA Kit be used with? Q1: Can an alternate buffer system be used with the IsoAmp tHDA Kit? A1: No, we would not suggest using a different buffer system. The supplied buffer has been optimized for HDA. Q2: Can HDA with the IsoAmp tHDA Kit be performed at temperatures other than 64°C? A2: Yes, 62°C - 65°C is the optimal temperature range for HDA. Q3: Can I perform HDA with the IsoAmp tHDA Kit in a thermocycler? A3: Yes, while HDA does not require a thermocycler, one can be used provided it is used at the suggested temperature. Q4: Can PCR primers be used with the IsoAmp tHDA Kit? A4: PCR primers can be used if they meet the selection criteria described in the Primer Design and Amplicon Selection section of the datacard. Q5: Can the HDA reaction using IsoAmp tHDA Kit be run in smaller reaction volumes (less than 50μl)? A5: Yes, reaction volumes from 25 - 50μl should perform similarly, provided the necessary adjustments are made to the protocol. Q6: Can the IsoAmp tHDA Kit amplify DNA sequences in excess of 120 bp? A6: No, HDA can amplify and detect short DNA sequences (70-120 bp) at a constant temperature. Q7: Can the IsoAmp tHDA Kit be used with Real-Time PCR machines? A7: HDA technology can work in real-time with the proper fluorescent dye on certain real time machines, however the IsoAmp tHDA Kit has not been optimized for this technique. Performance in real-time applications can not be guaranteed for all targets. Q8: Does HDA with the IsoAmp tHDA Kit require heat denaturation? A8: No, but a brief two minute heat denaturation may enhance performance. Q9: How much 100 mM MgSO4 should be added to the HDA reaction using the IsoAmp tHDA Kit? A9: We suggest performing a titration of 100 mM MgSO4 from 2.5 mM-5 mM to determine the optimal MgSO4 concentration for your primer set. Q10: What is the detection limit for the IsoAmp tHDA Kit? A10: As few as 10 copies of target DNA can be detected using optimized primers and buffers. Q11: What templates can the IsoAmp tHDA Kit be used with? A11: The IsoAmp tHDA Kit can be used with a variety of templates, including microbial genomic DNA, viral DNA, plasmid DNA and cDNA.