CRISPR-GENERATED CELL-BASED MODELS
When you can't find the right iPSC line or iPSC donor material to create a cell-based model for a specific disease, DefiniGEN can help. With our years of experience generating mature, primary-like cells from iPSCs and our proficiency at using CRISPR for targeted genome editing, we can build the cells that best reflect the biology you wish to model.
Want to do the differentiation yourself? We can perform the CRISPR genome editing and send you the bespoke iPSCs
Our custom cell model service is an integrated, end-to-end package that includes iPSC line or donor selection, the complete CRISPR editing workflow, and differentiation into the target cell type. From basic frameshift knock-out mutations to complex knock-in genetic changes, our dedicated scientific team will work collaboratively with you to design and build the most relevant cell-based model for your project.
Contact us to learn more about working with DefiniGEN - we respond within 1 business day to every inquiry.
*Using our in-house iPSCs can increase your chances of success as we already have a deep understanding of how these iPSCs will respond to CRISPR editing and during differentiation.
We can give your project a big head start with our well-characterized in-house iPSCs that model a number of monogenic diseases, many with multiple different models (Table 1 below).
When modelling any pathology using an iPSC differentiation platform, it is vital to confirm that the metabolic pathways implicated are expressed and active in the wild-type cells. DefiniGEN's hepatocytes are mature, predictive, and display all relevant biological pathways pertinent for our disease models, as these data show below.
|Acute intermittent porphyria|
|Genetic cholestasis (PFIC, TGP2, and Alagille syndrome)||Familial hypercholesterolemia|
|Wilson’s disease||Organic acidurias|
|Hereditary hemochromatosis||Cystic fibrosis|
|Tyrosinemia type 1||Erythropoietic protoporphyria|
|Alpha-1 antitrypsin deficiency||MCAD deficiency|
|Arginosuccinic aciduria (ASL)||D-bifunctional protein deficiency|
|Glycogen storage disease (GSD) type 1||Galactosemia Type 1|
|Urea cycle disorders||Citrullinemia|
|Crigler-Najjar syndrome||Familial amyloid polyneuropathy|
|Primary hyperoxaluria type 1||Atypical haemolytic uremic syndrome 1|
|Maple syrup urine disease (MSUD)|
The ATP7B gene is fundamental in hepatocytes for enabling the cells to effectively metabolise copper, and mutations in this gene can lead to Wilson’s disease. The ATP7B gene is shown here by qPCR to be expressed at comparable levels to human primary cells and the pathway is active in DefiniGEN’s differentiated hepatocytes, which make a good isogenic control for our disease model cells.
Gaucher’s disease is caused by the defective activity of the lysosomal hydrolase glucocerebrosidase, which is encoded by the GBA gene. The GBA pathway is present in DefiniGEN’s differentiated hepatocytes, shown here by qPCR, which provide a good isogenic control to our disease model cells.
Crigler-Najjar syndrome is a rare monogenic disorder which is caused by a mutation in the UGT1A1 gene. The UGT1A1 enzyme is required for the conjugation and excretion of bilirubin from the body. The UGT1A1 pathway is shown here by qPCR to be expressed sufficiently in DefiniGEN’s wild type differentiated hepatocytes for them to be used as isogenic controls to our disease model cells.
Familial Transthyretin Amyloidosis is a rare inherited condition characterized by abnormal build up of a protein called amyloid in the body. DefiniGEN hepatocytes present a good model system that mimic the disease phenotype and allow researchers to understand important disease-related mechanisms. This graph shows expression of the TTR pathway in our control wild type hepatocytes as comparable levels to human primary cells.
Alagille Syndrome is caused by mutations in the JAG1 and NOTCH2 genes. The NOTCH2 and JAG1 gene expression is shown here to be at comparable levels to human primary cells in DefiniGEN’s differentiated hepatocytes, and therefore provide a good isogenic control to our disease model cells.
The HFE gene is central in hepatocytes for enabling the cells to effectively metabolize iron, and mutations in this gene can lead to Haemochromatosis. The HFE pathway is shown here by qPCR to be expressed at sufficient levels in DefiniGEN’s differentiated hepatocytes, to provide a good isogenic control to our disease model cells.
It depends on the complexity of the project but generally we deliver the edited cell line in 8 weeks. If differentiation into a specific cell type is required, additional time will be necessary and will depend on the requested scale and cell type of interest.
We normally design up to 3 guide RNAs per model using software powered by the latest scoring algorithms. Depending on cutting efficiency (TIDE or ICE analysis), predicted on-target and off-target score we select the best guide for gene editing.
We work as a research partner offering collaborative services. We offer bi-weekly calls with our clients and interim emails with project reports. Our team is always available to provide technical support which can include experimental design and training on best practices for handling cells.
To generate a quote, we need your name, the name of your organization, your email address and details on the gene, mutation of interest, and cell type to edit. We also recommend contacting our team to go through project requirements in more detail. Contact us today >
We perform Sanger sequencing to show successful introduction of frameshift indels in the gene of interest. Additionally, we can evaluate protein expression via Western Blot or perform basic phenotypic screening.