Multiplex results obtained with one of the multiplex polymerase mix linked extremely well with the uniplex reference.
Amplifications were performed with a high-fidelity DNA polymerase mix.
All PCR was performed using Advantage 2 Polymerase Mix (Clontech, Mountain View, CA).
Qualitative RT-PCR assay was performed with Advantage 2 Polymerase Mix (BD Bioscience Clontech, Heidelberg, Germany).
Restriction enzymes, phosphatase, DNA polymerase mix, and T4 DNA Ligase were obtained from New England Biolabs (Ipswich, MA).
Neither DMSO nor betaine showed any marked influence on product formation using the stock polymerase mix for PCR when compared with the control samples (Fig. 2A and C).
10 µl of the reaction consisted of 1 µl cDNA, 0.8 µl MgCl2 (25 mM), 50 pmol of each primer, 1 µl polymerase mix and water.
Quantitative PCR shown in Figure 4 was performed with 1 µl cDNA in a 50 µl Platinum Taq Polymerase mix (Invitrogen) and in an exponential amplification condition (28 cycles) as determined in pilot experiments.
PCR reactions used the Advantage 2 Polymerase Mix (Clontech) under the following conditions: 94°C for 3 minutes, 24 cycles of 94° for 20 seconds, 68° for 6 minutes, followed by 1 cycle of 72° for 5 minutes.
Real-time PCR reactions were performed in triplicate using an ABI Prism 7000 system and SYBR green-Taq polymerase mix to determine the relative change in the expression of various KSHV genes.
The RT reactions were diluted 1 25, and 2 5 µl of diluted reaction was used for quantitative PCR using the 2× polymerase mix (Applied Biosystem) in a 25 µl reaction, with 1 µl of 5 mM of each primers.
