Stratagene’s Easy-A high fidelity PCR cloning enzyme is not just another polymerase. Like most scientists, I have used several different polymerases throughout my research career. To date, Easy-A polymerase is the best all-around enzyme I have found and is the primary polymerase I use in my research. Personally, I do a lot of cloning in my work and often need to amplify genes from cDNA. Since I want to clone genes with the correct sequence, I have tried several “high” fidelity enzymes (including Vent, Pfu and Pfu Ultra) but Easy-A has outperformed them all.
I have found that polymerases fall into two categories - those that only work well for amplification using large sources of template (plasmids) and those that are good for amplification using rarer templates (cDNA), as well. In my experience, most high fidelity enzymes fall into the first category: they are able to amplify from plasmids, but are unable to amplify from cDNA. In my work, I am rarely able to amplify a product from cDNA using Pfu and Pfu Ultra; however, these enzymes work very well when plasmid DNA is used as a template. In the past, I was forced to use Taq DNA polymerase (or variants of Taq that have higher fidelity, like Taq precision) to amplify from cDNA and could only hope to isolate a clone without errors. Of course, the larger the product, the higher the frequency of errors; hence, more clones were needed to find one with the correct sequence. Easy-A is the first high fidelity polymerase I have found that falls into the second category and has eliminated the need to use lower fidelity Taq variants to clone genes from cDNA. Using Easy-A, I have been able to amplify genes as large as 3.5 Kb from cDNA (so far I have not tried to amplify anything larger) and isolate a clone without any errors. Usually, I am able to isolate a perfect clone after sequencing just two or three clones.
Another advantage of the Easy-A polymerase is noted in its name. Most high fidelity enzymes cannot be used for TA cloning because they leave blunt ends. However, Easy-A polymerase adds the required 3’-A for TA cloning. This is extremely convenient since you can clone your PCR product without having perform restriction digestion first. In addition, Easy-A is a hot start polymerase which adds increased specificity. When compared to Taq DNA polymerase (or its variants Platinum Taq and Accuprime Taq) I have found that Easy-A generates comparable yields of PCR product.
One disadvantage of the Easy-A polymerase is the price, as it costs slightly more than generic Taq DNA polymerase. However, the cost compares well with other higher fidelity enzymes on the market. If you are looking for a robust enzyme and fidelity does not matter (for example, in typing) then I suggest sticking with Taq DNA polymerase, as the benefits of Easy-A are not needed. If you are looking for a robust enzyme that will give you high yields with increased fidelity and the capability to TA clone, then I recommend trying Stratagene’s Easy-A polymerase.
Aaron Tooley
Graduate Student
Dept. of Pathology
University of California, San Francisco