Long and complex PCR solutions: High-quality enzymes and mixes for complex PCR

ALLin™ Mega HS HiFi Polymerase - enzyme of choice for long and complex PCR applications

Product	Specialties	ALLin™ buffer with dNTPs	Fast cycling	GC/AT rich PCR	Sensitive, hot-start	Proofreading (3’-5’ exo)	Fidelity vs Taq	Max product	Direct PCR	Multiplex PCR	Cloning
ALLin™ Mega HS HiFi DNA Polymerase, 2 u/µl	Hot start version of ALLin™ Mega HiFi Polymerase: 100X higher fidelity than Taq for fast, complex, long PCR, excellent for NGS	●	●●	●●●	●	●●●	~100X	>20 kb	●	●●●	Blunt
ALLin™ Mega HS HiFi Mastermix, 2X	ALLin™ Mega HS HiFi Polymerase in 2X mastermix format	●	●●	●●●	●	●●●	~100X	>20 kb	●	●●●	Blunt
ALLin™ Mega HS HiFi Red Mastermix, 2X	ALLin™ Mega HS HiFi Polymerase in 2X ready-to-load red mastermix format	●	●●	●●●	●	●●●	~100X	>20 kb	●	●●●	Blunt
ALLin™ Mega HiFi DNA Polymerase, 2 u/µl	100X higher fidelity than Taq for a very fast, complex, long PCR, excellent for NGS	●	●●	●●●		●●●	~100X	>20 kb	●	●	Blunt
ALLin™ Mega HiFi Mastermix, 2X	ALLin™ Mega HiFi Polymerase in 2X mastermix format	●	●●	●●●		●●●	~100X	>20 kb	●	●	Blunt
ALLin™ Mega HiFi Red Mastermix, 2X	ALLin™ Mega HiFi Polymerase in 2X ready-to-load red mastermix format	●	●●	●●●		●●●	~100X	>20 kb	●	●	Blunt
ALLin™ HiFi DNA Polymerase, 2 u/µl	50X higher fidelity than Taq for Fast, GC rich, direct PCR with high fidelity	●	●	●●		●●	~50X	10 kb	●		Blunt
ALLin™ RPH Polymerase, 5 u/µl	Robust Proofreading Hot-start Polymerase for long hot start PCR	●	●	●●	●	●	~ 5X	35 kb	●●	●●	TA
ALLin™ RPH Mastermix, 2X	ALLin™ RPH Polymerase in 2X mastermix format	●	●	●●	●	●	~ 5X	35 kb	●●	●●	TA

Long/Complex PCR and PCR Troubleshooting

Do you have problems with PCR yield? Reach out to PCR troubleshooting experts at info@highqu.com for personalized assistance with PCR troubleshooting. Most common reasons of PCR failures can be:

If there is a non-specific amplification or PCR background:

• Ensure that the primers are designed correctly by using primer design tools.

• Adjust the primer concentration to favour specific primer annealing over non-specific binding.

• Run an annealing temperature gradient to find the optimal temperature for each new template-primer-PCR mix set. HighQu mixes “like” higher annealing temperatures.

• Reduce the template amount, and if possible, check the template DNA integrity on the agarose gel, it can be that your gDNA sample is degraded.

• If nothing helps, switch to hot-start PCR to prevent non-specific amplification during the initial stages of PCR troubleshooting.

If there is low PCR product yield or no amplification:

• In addition to told above, try to reduce template amount, especially when working with crude samples, you may have PCR inhibitory effects coming from impurities.

• Evaluate PCR cycling conditions: assess the denaturation, annealing, and extension times to ensure sufficient amplification, especially for longer targets.

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