PrimerBlast was developed by NCBI and combines the tried and true Primer3 platform with Blast capabilities. NCBI’s PrimerBlast (Primer3): Possibly one of the most highly regarded and versatile online tools for primer design and analysis. Other considerations: Not high throughput (sequences are analyzed one at a time manually at the time of writing).Ģ.Good for users who: Are new to primer design, know what target range for GC content and melting temperatures they want, and are amplifying DNA from a plasmid for typical cloning experiments.Since the added site doesn’t bind to template DNA in the first round of amplification, you should ensure that the primer properties do not change significantly and inhibit the reaction! I find the hairpin structure formation tab particularly fun, and the latest version also allows users to perform an NCBI Blast easily without copying and pasting the sequence.Ī word to the wise: check on the primer without any added restriction sites for cloning before ordering your primer. After plugging in a sequence, you will get information on sequence length, GC content, melting temperature range, molecular weight, extinction coefficient, and optical density. IDT OligoAnalyzer: This is a simple yet effective tool for determining physical properties of oligo sequences up to 255 bases. And after that, check out the e-book “ The Bitesize Bio Guide to PCR” to become a master of this invaluable molecular biology technique.ġ. Below are four of my favorite online programs available to make primer design quick, easy, and effective.Ī quick note before we get started: If you’re new to PCR, Bitesize Bio has a plethora of articles for you! Learn how polymerase chain reaction works, what to do when it doesn’t work, and variations on standard PCR. Designing and running PCR reactions in the lab has become so commonplace that the number of primer design tools available can be a bit overwhelming for a beginner (or even an experienced molecular biologist!).
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