Codon Optimization

To maintain wild-type protein expression:

• We avoid using rare codons (those with a codon frequency of <8%).
• We ensure sequences do not have strong hairpins (ΔG < -8) in the first 48 base pairs of resulting sequences.
• We check both strands to ensure they don’t contain the enzyme cut sites you asked us to avoid.
• We avoid introducing promoter sequences internal to expression sequences by avoiding the creation of strong sigma70 binding sites.
• We avoid sequences that create strong ribosome binding sites (GGAGG and TAAGGAG).
• We avoid sequences that create terminator sequences (TTTTT or AAAAA).

To improve the likelihood of successful manufacture:

• We won’t introduce any repeat longer than 20 base pairs.
• We avoid homopolymer runs of 10 or more bases.
• We avoid fitted sequences that create global GC% of less than 25% or more than 65% and local GC windows (50 bp) of less than 35% or more than 65%.

Please note that codon optimization is useful for improving chances of a successful synthesis, not protein expression.  

Currently, we are unable to synthesize genes that score as Impossible. There are several factors that cause our algorithm to score a gene Impossible and the specific parameters are unique to the gene sequence. Our algorithm is proprietary and was developed using machine learning. In addition to machine learning, our algorithm does take into consideration the following factors:

• %GC between 25-65%
• Max homopolymer length < 10
• Low homology

A unique combination of the above is a reflection of the specific gene sequence score of Impossible.

Here is a list of organisms that are supported for codon optimization on our website:

• Arabidopsis thaliana
• Aspergillus niger
• Aspergillus oryzae
• Bacillus subtilis
• Brassica napus
• Caenorhabditis elegans
• Cricetulus griseus (CHO)
• Drosophila melanogaster
• Escherichia coli
• Glycine max
• Homo sapiens
• Hypocrea jecorina
• Medicago sativa
• Mus musculus
• Nicotiana tabacum
• Petunia x hybrida
• Pichia pastoris
• Pisum sativum
• Rattus norvegicus
• Saccharomyces cerevisiae
• Solanum tuberosum
• Spodoptera frugiperda
• Streptomyces coelicolor A3(2)
• Sus scrofa
• Toxoplasma gondii
• Xenopus laevis
• Yarrowia lipolytica
• Zea mays

If you do not see your specific organism, contact customersupport@twistbioscience.com for help. 

To maintain wild-type protein expression:

• We avoid using rare codons (those with a codon frequency of <8%).
• We ensure sequences do not have strong hairpins (ΔG < -8) in the first 48 base pairs of resulting sequences.
• We check both strands to ensure they don’t contain the enzyme cut sites you asked us to avoid.
• We avoid introducing promoter sequences internal to expression sequences by avoiding the creation of strong sigma70 binding sites.
• We avoid sequences that create strong ribosome binding sites (GGAGG and TAAGGAG).
• We avoid sequences that create terminator sequences (TTTTT or AAAAA).

To improve the likelihood of successful manufacture:

• We won’t introduce any repeat longer than 20 base pairs.
• We avoid homopolymer runs of 10 or more bases.
• We avoid fitted sequences that create global GC% of less than 25% or more than 65% and local GC windows (50 bp) of less than 35% or more than 65%.

Please note that codon optimization is useful for improving chances of a successful synthesis, not protein expression.  

Currently, we are unable to synthesize genes that score as Impossible. There are several factors that cause our algorithm to score a gene Impossible and the specific parameters are unique to the gene sequence. Our algorithm is proprietary and was developed using machine learning. In addition to machine learning, our algorithm does take into consideration the following factors:

• %GC between 25-65%
• Max homopolymer length < 10
• Low homology

A unique combination of the above is a reflection of the specific gene sequence score of Impossible.

Here is a list of organisms that are supported for codon optimization on our website:

• Arabidopsis thaliana
• Aspergillus niger
• Aspergillus oryzae
• Bacillus subtilis
• Brassica napus
• Caenorhabditis elegans
• Cricetulus griseus (CHO)
• Drosophila melanogaster
• Escherichia coli
• Glycine max
• Homo sapiens
• Hypocrea jecorina
• Medicago sativa
• Mus musculus
• Nicotiana tabacum
• Petunia x hybrida
• Pichia pastoris
• Pisum sativum
• Rattus norvegicus
• Saccharomyces cerevisiae
• Solanum tuberosum
• Spodoptera frugiperda
• Streptomyces coelicolor A3(2)
• Sus scrofa
• Toxoplasma gondii
• Xenopus laevis
• Yarrowia lipolytica
• Zea mays

If you do not see your specific organism, contact customersupport@twistbioscience.com for help.