GeneGenie: optimized oligomer design for directed evolution

Nucleic Acids Res. 2014 Jul;42(Web Server issue):W395-400. doi: 10.1093/nar/gku336. Epub 2014 Apr 29.


GeneGenie, a new online tool available at, is introduced to support the design and self-assembly of synthetic genes and constructs. GeneGenie allows for the design of oligonucleotide cohorts encoding the gene sequence optimized for expression in any suitable host through an intuitive, easy-to-use web interface. The tool ensures consistent oligomer overlapping melting temperatures, minimizes the likelihood of misannealing, optimizes codon usage for expression in a selected host, allows for specification of forward and reverse cloning sequences (for downstream ligation) and also provides support for mutagenesis or directed evolution studies. Directed evolution studies are enabled through the construction of variant libraries via the optional specification of 'variant codons', containing mixtures of bases, at any position. For example, specifying the variant codon TNT (where N is any nucleotide) will generate an equimolar mixture of the codons TAT, TCT, TGT and TTT at that position, encoding a mixture of the amino acids Tyr, Ser, Cys and Phe. This facility is demonstrated through the use of GeneGenie to develop and synthesize a library of enhanced green fluorescent protein variants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Algorithms
  • Codon
  • Directed Molecular Evolution / methods*
  • Genes, Synthetic*
  • Green Fluorescent Proteins / chemistry
  • Green Fluorescent Proteins / genetics
  • Internet
  • Oligonucleotides / chemistry*
  • Sequence Analysis, Protein
  • Software*


  • Codon
  • Oligonucleotides
  • enhanced green fluorescent protein
  • Green Fluorescent Proteins