Design all 5861 edits covering the entire 292 AA sequence of dapA in less than 10 minutes.
Insert a promoter sequence in front of 4327 genes using 3 different strategies — all in less than 35 minutes.
See how you can design a KO library covering 95% of the yeast genes in less than an hour.
Still wondering what you can do with the Onyx? The table below lists a sampling of the libraries you can easily build using the Onyx Platform. Click the name of the library to see a description of the library and how to use them.
(AA: amino acid, nt: nucleotide)
Target type: Single location
Purpose: This approach enables the discovery of novel variants that improve enzyme function. Using this unbiased strategy can uncover beneficial diversity in unexpected sites in a protein.
Example libraries:
See how to design a library in InscriptaDesigner® software: dapA full saturation library design (2:49 min)
See examples of a project using this library:
Target type: Single location, pathway
Purpose: A hybrid between purely arational full AA saturation (20 codons tested per AA position) and targeted AA substitutions, this approach designs a specific reduced set of AA substitutions at every position and/or targets only a reduced subset of positions.
Example of libraries:
See examples of a project using this library:
Target type: Single location, pathway
Purpose: Keys residues or “hot-spots” in multiple genes can be discovered in a single experiment. Minimizing the number of Onyx edits per residue can allow you to target many genes in one experiment, and subsequent libraries can do full AA saturation on key regions.
Example of libraries:
Target type: Single location, pathway
Purpose: Hypothesis-guided engineering can lead to an increase in the frequency of hits. Testing multiple hypothesis-guided strategies could also lead to learnings that inform future campaigns, increasing the efficiency of forward engineering over time.
Specific AA substitutions could be chosen based on: structure-guided engineering strategies, evolutionary strategies guided by multiple-sequence alignments, prior datasets identifying key domains or key residues, etc.
Example of libraries:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Altering global regulation through post-translational modifications can cause larger impacts on cell physiology with a smaller number of more targeted edits.
Example of libraries:
Target type: Single location
Purpose: In forward engineering, a winning strategy is often to find beneficial variants and then consolidate those variants into a single strain to find combinations that yield further phenotypic improvements. A single Onyx edit is targeted to one site in the genome, but within that Onyx edit, multiple changes to the genome can be made at one time. For example, making 6 different, targeted AA substitutions within a 15 AA window in a gene.
Example of libraries:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Localization of enzymes plays a key role in their biological function. Disrupting or “re-wiring” localization within the cell can cause significant biological changes which may be beneficial to desired phenotypes.
Example of libraries: Modify the secretory machinery of the cell by changing the localization of key enzymes to different cellular compartments. For all proteins known to be involved in secretion, replace any endogenous localization signals with an NLS (sequester in the nucleus) and Golgi localization signal.
Target type: Single location, pathway
Purpose: Evolution has found many “solutions” to biological problems already, but it can be challenging to synthesize and screen all known orthologs of a single enzyme. With an Onyx library, you can deliver a mix of different variants that are found in nature to a single base enzyme expressed from the genome.
Example of libraries:
Target type: Pathway
Purpose: If you have a protein pathway that you want to purify, instead of tagging each protein separately in different strains, you could introduce His-tags to all of those proteins at one time in a pooled format, and then purify them all together in a “one-pot” purification.
Example of libraries:
Target type: Pathway, genome-wide
Purpose: Many proteins have separate functional domains, and their activity/stability/function can be modulated with domain truncations that would not be possible with full gene knock-outs.
Example of libraries:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Many proteins have separate functional domains, and their activity/stability/function can be modulated with domain truncations that would not be possible with full gene knock-outs. The N‑terminus is also a common site for signal and degradation signals and other regions of high biological significance to the protein’s function.
Example of libraries:
Target type: Single location, pathway
Purpose: Expression tuning can be critical for metabolic engineering and many other applications — often, even just small, localized changes in the promoter region can have large impacts on gene expression.
Example of libraries:
Target type: Single location, pathway
Purpose: Higher fold-changes in expression can often be achieved by combining nt substitutions into combinatorial Onyx edits in the promoter region. 1000s of combinations of known beneficial edits, or combinations of variants found in divergent regions of the promoter can be tested in a single Onyx library.
Example of libraries:
Target type: Single location, pathway
Purpose: Translation rate is often determined by the sequence adjacent to the start codon — by making synonymous substitutions at the beginning of a gene, expression can be modulated.
Example of libraries:
Target type: Pathway
Purpose: Synthetic promoters and/or RBS elements can be used to tune the expression of genes. When targeting a pathway or set of functionally-related genes, a “ladder” of synthetic promoters of known strength can be used to find the ideal expression level in a single experiment.
Example of libraries:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Translation initiation efficiency impacts protein levels and can be regulated by editing the nucleotide sequence adjacent to the start codon.
Example of libraries:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Alternative start codons like GTG can change expression with a very small edit that can be readily performed across pathways or genome-wide.
Example of libraries:
Target type: Pathway
Purpose: Codon usage can impact the expression of a gene (or of adjacent genes!). Targeted codon optimization or de-optimization can be a way to modulate expression without changing enzyme activity.
Example of libraries:
See examples of a project using this library:
Target type: Pathway
Purpose: A variety of different Onyx edits (synthetic promoter “ladder”, KO, RBS, UTR deletions, etc.) can be combined in a single library to modulate the expression of each gene in multiple ways within the same experiment. This can be very useful in identifying genes where phenotype increases or decreases as the expression is increased.
Example of libraries:
Target type: Pathway, genome-wide
Purpose: Targeted codon changes based on a prediction of optimal yeast codon usage in all key enzymes in an engineered heterologous pathway.
Example of libraries:
See how to design a library with InscriptaDesigner™ software:
See examples of a project using this library:
Target type: Pathway, genome-wide
Purpose: Synthetic promoters or terminators can lead to strong over/under-expression of genes relative to their endogenous regulation, enabling unbiased pathway-wide or genome-wide surveys to find key genes involved in a phenotype of interest.
Example of libraries:
Target type: Pathway, genome-wide
Purpose: Transcripts stability can play a big role in expression level — deletions of the UTR region(s) tend to decrease expression levels, which can help balance pathway flux or identify novel biological processes of interest.
Example of libraries:
Target type: Pathway, genome-wide
Purpose: Gene regulation networks are ideal targets for genome engineering — a single Onyx edit in a cell can lead to key changes across a network of genes. Onyx libraries can target transcription factor genes with edits that increase and/or decrease expression, and they can also target the transcription factor binding sites (TFBS) that are regulating the expression of individual target genes.
Example of libraries:
Target type: Single location, pathway
Purpose: ncRNAs play various roles in the eukaryotes — Onyx edits can precisely target these non-coding features to try to modulate their role in gene regulation, RNA metabolism, etc.
Example of libraries:
Target type: Single location, pathway
Purpose: Transposons and other mobile elements can affect the local heterochromatin state and genome stability. Onyx edits can remove certain regions of these features to impact the expression and stability of nearby regions.
Example of libraries:
Target type: Pathway, genome-wide
Purpose: ncRNAs play various roles in the eukaryotes — Genome-wide Onyx edits can disrupt the regulatory and other functions of these ncRNAs by creating deletions that disrupt their function.
Example of libraries:
Target type: Single location, pathway, genome-wide
Purpose: A heterochromatin state is a critical way to regulate gene expression, and heterochromatin “spreading” can have far-reaching consequences on the regulation of many nearby genes. Many sequence motifs and elements are known to impact chromatin state (tRNA genes, TAD boundaries, nucleosome positioning motifs, etc).
Example of libraries: