MX3000P qPCR Machine From Stratagene

Real-time quantitative PCR (qPCR) is one of the latest modifications of PCR that has greatly enhanced gene expression research. One major step in validating microarray experiments has been the amplification of a select set of genes shown in the microarray to be of some importance. qPCR has also been useful in elucidating the expression level in various genes of interest when these genes are already known to play a part in the biological system studied.

Stratagene developed a product that will fit easily into many labs’ day-to-day experiments. The MX3000P qPCR machine is a great value and comes with easy-to-use fully integrated software, MX Pro™. Once the machine is up and running, the user-friendly software will help make developing protocols and interpreting data straightforward. The software, MX Pro, is well integrated with the machine and experimental design is straightforward requiring only a few mouse clicks to determine which wells contain which dyes, and labels for each well. Once you start the experiment, a real-time graph of your data is presented to you, and can be a very useful tool in troubleshooting. I also found the analysis part of the software to be very useful in quickly interpreting data. I have performed both relative gene expression and absolute gene expression and have found the software easy to display both types of experiments in a easily understandable format. One of the best features of the software is the custom reports that the program will provide. In seconds, you can have a great looking report including amplification graphs, standard curves, thermal profile, and anything else you could need for your run. The machine has also been designed to work with all fluorescent chemistries used at this time (SYBR Green, TaqMan® Probes, Scorpion Probes, etc.).

Unfortunately, qPCR itself is not an easily mastered technique. A common strategy for new labs starting qPCR is to use SYBR Green: a double-stranded DNA staining fluorescent dye, to quickly obtain data from a variety of already available primer sets. SYBR Green has its drawbacks as it is known to inhibit Taq polymerase and thus, can result in less then the optimal doubling of template per cycle. Although not a problem for highly expressed genes, low expression genes may not be detectable in this manner. To overcome this drawback, multiplexing can be attempted after optimizing individual primer sets with a fluorescently label probe. This approach shows some of the drawbacks to this entry-level machine. Running an internal dye standard (which is recommended but not required) results in having only 3 fluorescent colors to design experiments with. Other, more expensive, instruments offer more filter set options.

I have used this machine for quantification of various genes using SYBR Green and I have found it to be a very reliable and dependable machine. I have also used this experiment to test multiple gene products using multiplexing with Taqman® probes. These experiments are more complex then SYBR green reactions, and required more wells for various controls, such as each primer set and probe alone. These studies using multiplexing revealed some of the machines shortcomings, such as the 96 well limit, but overall this machine is a great value. In most labs, these shortcomings would go unnoticed, but if using extensive fluorescent chemistry used, I would recommend considering a more robust machine.