Flexi® Vector Primer Design Tool

The Flexi® Vector Systems use two rare-cutting restriction enzymes, Sgf I and Pme I, in a simple, yet powerful, directional cloning method for protein-coding sequences. These systems provide a rapid, efficient and high-fidelity way to transfer these sequences between a variety of Flexi Vectors. The Flexi Vectors contain various expression or peptide tag options to enable expression of native or fusion proteins to study protein structure and function, as well as protein-protein interactions.

This software will design PCR primers for use with Flexi® Vectors which will amplify a coding region from your input sequence. These primers will have the following characteristics

• The primer pair will have
  • A theoretical Tm between specified limits
  • Minimal primer dimer potential
  • Minimal self-annealing or hairpin potential
• The 5' primer will have (from 5' to 3')
  • 4 arbitrary bases
  • the sequence GCGATCGCCATG which has a SgfI site
  • An ATG start codon in frame with your reading frame if desired/required.
  • 20-35 bases of homology with the 5' end of your input sequence
• The 3' primer will amplify the sequence up to or including the first detected in-frame stop codon (TAA, TAG or TGA) and will have (from 5' to 3')
  • 4 arbitrary bases
  • the sequence GTTTAAAC, which contains the PmeI site encoding a Valine (GTT) and a stop codon (TAA)
  • 20-35 bases of homology with the 3' end of your input sequence

Go directly to the FlexiVector Primer Design Tool next time.