ApE Software

You can download ApE, a plasmid editing software, here.

ApE is used to view and edit potential and current DNA sequences (such as plasmids, g-blocks etc.)

How to view the sequence of a feature (primer, tag, etc.):
At the top of the page is a list of features of your current sequence. To highlight where each feature is in your DNA sequence, double click on the name of the feature you want to view in the list. Additionally, if you are moving your mouse over a certain sequence, the name of the features in that area will appear underneath the list of features at the top. This will help you to determine where one feature ends, and another begins, or where they overlap.

How to add a new feature:
Highlight the sequence in the file, and right click and select add new feature. (Alternatively, you can highlight the sequence and go to the dropdown menu at the top of the page to add a feature.) After adding a feature, a new window will pop up. Change the color to something different than the surrounding colors, and make sure to name it something descriptive. (Note- DO NOT select reverse complement! This will change the actual sequence to the reverse complement and cause all sorts of problems!)

How to determine Tm:
PCR usually requires that you know your Tm, or the melting temperature of the PCR product. For best results when designing primers, try to get the Tm's of primes in the same reaction within 5 degrees of one another, for best results. Usually, you'll pick the lowest Tm and subtract 5 degrees for best results with PCR amplification

With Gibson- Usually, with PCR for Gibson, you will design primers that extend farther than your DNA, to create overlaps between two sequences for Gibson. Therfore, when determining Tm for PCR, you will need to highlight the piece of the primer that will actually bind to the DNA, and exclude the overhang. At the top of ApE in the right corner will be "Tm= " which will tell you your correct Tm.

With general PCR- Simply highlight the entire primer and read the "Tm= " in the top right corner.