A library preparation kit for Next Generation Sequencing using an Illumina® platform. This assay detects the Onyx edit barcode, allowing users to derive the distribution of edits within the cell population.
A library preparation kit for Whole Genome Sequencing using an Illumina™ platform. This assay detects the edit sequence in the edited cell genome and can be run either on isolates (to identify the edit present in a particular clonal isolate) or pooled samples (to measure the overall edit fraction of cells).
The Onyx Genotyping Kits make it easy to characterize and asses your genome engineering runs prior to phenotyping. Now you can confidently plan your downstream phenotyping experiments.
The Onyx Barcode Diversity Assay
Edit Identification Assay
Phenotyping is typically done in one of two ways, isolated or pooled. Determining which strategy to use depends on the goals of the experiment. Learn more about the phenotyping workflows utilized in Inscripta’s lab — read our phenotyping technical note.
Step 1: Isolate phenotyping
Onyx cell library clones are isolated and phenotyped individually. The number of clones screened to identify a certain number of edits is determined using the screener’s curve.
Learn about Inscripta’s Lysine production strain engineering application which resulted in a 14,000-fold increase of lysine production in E. coli.
Step 2: Identify the edits of interest
Edited cells with the phenotype of interest are then genotyped using the edit identification assay to determine the edit responsible for the phenotype.
Step 1: Pooled phenotyping
Pooled cells are grown in the presence of some selection pressure (drug, stress factor, etc.). Both positive and negative selection schemes can be employed to identify edits that respond to the agent being tested. The selector score is used to determine the number of cells to seed a selection experiment with, to ensure sufficient library representation.
Download the application note for an example of a pooled phenotyping study using the Onyx Platform.
Step 2: Identify the edits of interest
The cell population is then sequenced using the barcode diversity assay. This allows one to determine which edits are enriched or depleted and by extension deemed to provide a beneficial or deleterious contribution to cell growth.