The ADA Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population generated from the near-haploid human HAP1 cell line, designed to disrupt the endogenous ADA gene. This product provides a loss-of-function model for adenosine deaminase, enabling investigations into purine metabolism, adenosine signaling, and immune cell development.
The HAP1 cell line is a near-haploid derivative of the KBM-7 chronic myeloid leukemia line, exhibiting a largely haploid karyotype and adherent growth. Its haploid genome simplifies knockout studies because disruption of a single allele can yield a functional null phenotype, making it a powerful model for functional genomics. HAP1 retains a hematopoietic gene expression program, rendering it suitable for immune-related investigations.
ADA catalyzes the deamination of adenosine and 2??-deoxyadenosine, a critical step in purine catabolism that prevents accumulation of dATP, a potent inhibitor of ribonucleotide reductase and DNA synthesis. ADA expression is regulated by T-cell receptor signaling through NF-??B and AP-1, and is further stimulated by IL-2. The enzyme interacts with DPP4/CD26 and the cofactor ADCP2. Its activity modulates signaling through adenosine receptors ADORA1 and ADORA2A, and influences downstream purine nucleoside phosphorylase and DNA synthesis pathways. Loss-of-function mutations in ADA lead to ADA-SCID, characterized by severe lymphotoxicity from unmetabolized purine nucleosides.
In the haploid HAP1 background, ADA knockout provides a clean genetic model to dissect the metabolic and signaling consequences of adenosine accumulation without the confounding effects of a second wild-type allele. The polyclonal nature of this product offers a pooled representation of multiple CRISPR-induced mutations, allowing researchers to assess average phenotypic responses that are less susceptible to clone-specific artifacts. This model is especially valuable for studying the mechanisms of purine nucleoside toxicity in a human immune-competent context, recapitulating key aspects of ADA-SCID pathophysiology at the cellular level.
Typical applications include modeling ADA-SCID and purine metabolism disorders, functional genomics screens to discover ADA genetic interactions, and drug screening for immunomodulators. These cells also serve as a platform for evaluating adenosine analogs and validating gene therapy targets. Relevant assays include HPLC-based metabolite profiling, proliferation and apoptosis measurements (MTT, Annexin V), Western blotting and RT?qPCR for ADA expression, cAMP assays for adenosine receptor activity, and flow cytometry for DPP4. For further details, contact Ascent Research.
