Addition of pcr reagents to master mix - What order is best? (Dec/19/2005 )
What is the best order in which to add reagents to make the PCR master mix? I have been adding them in this way:
- 1. Buffer
- 2. MgCl2
- 3. dNTP's
- 4. AmpErase UNG
- 5. DNA Poly
I just read, however, that the MM should be dispersed into tubes, followed by MgCl2 and template.
Other than making sure not to add enzyme to a mix without buffer, does the order make a difference? Sorry if this question is very naive!!
4. AmpErase UNG
aliquot out resulting MM.
Really shouldn't matter that much, though...