GEN Protocols

Genome Editing

Protocol

Lentiviral titering by crystal violet staining

This protocol provides an example for titering lentiviral particles using crystal violet staining in a 6-well format to obtain transducing units per mL (TU/mL). Once lentiviral particles are packaged and collected, small aliquots should be frozen at –80°C. One aliquot should be thawed on ice and use... More

Protocol

Multiplexed, trackable CRISPR-based genome engineering for optimization of heterologous protein expression and activity in E. coli

This work demonstrates the use of the Onyx platform for interrogation and optimization of heterologous proteins in E. coli. Compared to traditional methods for protein engineering, such as random mutagenesis or targeted site-directed mutagenesis approaches, the Onyx technology dramatically increases... More

Protocol

Massively parallel genome engineering followed by pooled growth selections for rapid target discovery in microbes

The ability to engineer the entire genome of microorganisms at a massively parallel scale using the Onyx platform will reshape the manner in which researchers perform pooled cultivation experiments (such as ALE) and generate data sets. Here, Onyx technology was used to rapidly generate genomewide k... More

Protocol

Dharmacon Edit-R crRNA Libraries

Libraries of predesigned synthetic crRNAs provide an opportunity to apply the CRISPR-Cas9 system for functional gene knockout analysis, or dCas9-VPR system for transcriptional gene activation, in an arrayed format. Dharmacon crRNA libraries for functional gene knockout consist of Edit-R synthetic cr... More

Application

Rebuilding evolutionary success: Using Inscripta’s Onyx Digital Genome Engineering Platform to ease and enhance Adaptive Laboratory Evolution experiments

Adaptive Laboratory Evolution (ALE) is a powerful approach to leverage biology’s evolutionary capacity to develop strains that thrive in challenging growth environments, with applications to biotechnology and bioproduction and for basic biochemical discovery. ALE experiments have been enabled by nex... More

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