Conventional PCR Amplification
Conventional (end-point) polymerase chain reaction (PCR) enables amplification of specific DNA fragments with the help of a thermocycler and is one of the most widespread molecular biology techniques, with endless applications spanning molecular cloning to genotyping, to semi-quantitative gene expression analysis to molecular diagnostics.
End-Point PCR is distinguishable from real-time PCR as it is not quantitative; the results are instead visualized on a gel once the run has finished.
There are of course a variety of kits available on the market that are providing all the necessary PCR reagents for a successful End-Point PCR reaction. When choosing a kit, there are a few things to consider when picking the one for your experiments:
The first is templates, as DNA and RNA require different procedures. For example, if you are performing Reverse-Transcription PCR (RT-PCR), an RNA template is required to produce complementary DNA (cDNA). Such RT-PCR procedures are often divided into one- or two-step PCRs.
Next, choosing the right polymerase can be crucial for a successful experimental outcome.
Polymerases may differ in their processivity. The processivity is defined as the number of bases they will read on a DNA strand before falling off the template. This is important for cloning of long fragments to avoid truncated amplicons.
Some polymerases can also have proofreading activity, like the famous Klenow polymerase, which ensures that amplified fragments are a high fidelity copy of the original. This is generally measured as average mistakes per bases amplified and should be kept as low as possible to avoid unexpected mutations particularly during cloning experiments. If high fidelity is not essential (like for confirming plasmid inserts), then Taq polymerase is a good, cost-effective alternative.
Hot start PCR compatible polymerases can also help you to get rid of unwanted non-specific amplification.
The polymerase processing speed is another factor to consider. High-speed polymerases can save you time when using PCR programs with a long primer extension cycles.
Complex applications such as multiplex PCR, where the amplification of multiple targets in the same reaction occurs, or the creation of labeled DNA probes using modified dNTPs, also have dedicated kits to make your experimentation easier.
Our End-Point PCR Troubleshoot will simplify the process further.
Compare Reverse Transcription End-Point PCR Kits
With our compare function, you can avoid all the time and energy wasted sifting through multiple web pages from different suppliers. At ZAGENO you can clearly see kits side-by-side, with the relevant attributes for each kit neatly in line for easy selection of the best product for you.
The ZAGENO comparison does not highlight one kit to be better than the other, as the kit of choice may vary between researchers - depending on each individual's preferred attributes. The best kit is the one that meets your needs - ZAGENO allows you to make an informed decision with minimum effort.
Check out our How It Works page for a guide to using the comparison function.
Video: Oxford Academic (Oxford University Press)/YouTube