( the next page. For maximum yield and s...)HotStarTaq PCR Handbook

the next page. For maximum yield and specificity, temperatures and cycling times should be optimized for each new template target or primer pair.

7. Place the PCR tubes in the thermal cycler and start the cycling program.
Note: After amplification, samples can be stored overnight at 28C or at 20C for longer storage.

PCR Protocol Using HotStarTaq Master Mix

This protocol serves only as a guideline for PCR amplification. Optimal reaction conditions, such as incubation times and temperatures, and amount of template DNA, may vary and need to be determined individually.

Notes:
Each PCR program should be started with an initial activation step of 15 min at 95C to activate HotStarTaq DNA Polymerase (see step 6 of this protocol).
HotStarTaq Master Mix provides a final concentration of 1.5 mM MgCl2 in the final reaction mix, which will produce satisfactory results in most cases.
However, if a higher Mg²+ concentration is required, prepare a stock solution containing 25 mM MgCl2.
Set up reaction mixtures in an area separate from that used for DNA preparation or PCR product analysis.
Use disposable tips containing hydrophobic filters to minimize crosscontamination.

1. Thaw primer solutions.
Mix well before use.
Optional: prepare a primer mix of an appropriate concentration (see Table 4) using the water provided.<
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