Oligo Modifications
Support and Educational Content

Generate codon balanced libraries for mutagenesis with trimer modifications

Avoid stop codons and limit amino acid redundancy with Trimer 19 or Trimer 20 codon modifications

Quick facts:

Availability: DNA

Location: 5', internal, and 3'

Scales: 100 nmol to large scale

Purification: Standard desalt or HPLC

Ordering:

  • Trimer 19 modification code /iTriMix19/

  • Trimer 20 modification code /iTriMix20/

Please email noncat@idtdna.com for custom trimer mix requests.

Optimizing oligonucleotide libraries for mutagenesis

Oligonucleotide libraries are commonly used for mutagenesis of proteins that can be screened for potential improvements in biological function, such as enhanced expression levels, stability, solubility, enzymatic activity, or binding affinity. While you can create libraries of sequences by simply having random bases incorporated into synthetic oligonucleotides, this method introduces stop codons and a higher proportion of certain amino acids, which increases the time and resources required to screen the resulting libraries. Alternatively, using the 3 degenerate bases NNK will reduce, but not eliminate the number of possible stop codons and improves proportional codon biases, but will not eliminate, the number of possible stop codons and codon biases.

These issues can be avoided by incorporating codon trimers into your oligos. IDT trimer mixes do not contain stop codons, and trimer mixes provide a balanced mix of the amino acid codons, resulting in equal representation of each unique codon.

Trimer codon mix options for balanced amino acid coding

IDT provides 2 trimer codon mixes, Trimer 19 Mix and Trimer 20 Mix, that can be incorporated into IDT oligos.

Trimer 19 contains an equal ratio of the following 19 codons (excluding CYS):

  • AAA, AAC, ACT, ATC, ATG

  • CAG, CAT, CCG, CGT, CTG

  • GAA, GAC, GCT, GGT, GTT

  • TAC, TCT, TGG, TTC

Trimer 20 contains an equal ratio of the following 20 codons (including CYS):

  • AAA, AAC, ACT, ATC, ATG

  • CAG, CAT, CCG, CGT, CTG

  • GAA, GAC, GCT, GGT, GTT

  • TAC, TCT, TGC, TGG, TTC

To limit synthesis complexity and facilitate fast delivery, we restrict the total length of oligos with trimer insertions to 100 total bases (counting each trimer as 3 bases). The number of codon trimer insertions within an oligonucleotide is also limited to 15. However, we are able to consider other designs outside of these specifications as non-catalog requests. Just contact us at noncat@idtdna.com.

Create your own custom trimer codon mixes

We can also incorporate a custom trimer codon into oligos. This allows you to customize the ratios of trimers incorporated. To order a custom trimer codon mix, email noncat@idtdna.com with your name, organization, and sequence designs. Oligos containing Custom Trimer Mixes can be ordered as standard desalted oligos (recommended for oligos containing mixes), provided that other modifications included in the sequences do not require HPLC.

Further reading

Oligonucleotide modifications: Choosing the right mod for your needs—Review these guidelines on selecting oligonucleotide modifications to learn how they can help you in your research.

Need a non-standard modification?—Did you know that IDT offers 89 modifications that are not listed in our online catalog? A few of the more popular ones are described along with information on how to order them. IDT will consider any modification you require. Please make such inquiries to noncat@idtdna.com.

Need a library of related DNA or RNA oligo sequences?—Build variability into your oligo sequences by incorporating Mixed Bases. We offer mixes of multiple base types as well as nonstandard and modified bases.

Increase the Tm of short, AT-rich primers and probes—Add this modified base to increase the melting temperature (Tm) of primers and probes. It is especially useful when you need to work with short, A-T rich sequences.

Inverted bases—Learn how inverted bases allow you to reverse the orientation of part of your oligo sequence or add a 5'-end restricted modification to the 3'-end of your sequence.


Author: Stephanie Youtsey is the non-catalog coordinator in the Specialty & Manufacturing Services group at Integrated DNA Technologies.

© 2016 Integrated DNA Technologies. All rights reserved. Trademarks contained herein are the property of Integrated DNA Technologies, Inc. or their respective owners. For specific trademark and licensing information, see www.idtdna.com/trademarks.


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