MFSD2A Knockout HCT116 Cell Line

The choice depends on whether you are studying MFSD2A (major facilitator superfamily domain containing 2A)'s role as the principal blood-brain barrier (BBB) DHA (docosahexaenoic acid) transporter or modeling autosomal recessive primary microcephaly type 15. The Knockout line is the standard tool for asking whether MFSD2A is required for sodium-dependent uptake of lysophosphatidylcholine (LPC)-DHA and other omega-3 LPCs across the BBB — MFSD2A is selectively expressed on the luminal endothelial membrane of brain microvessels and is the major transporter for delivering essential omega-3 fatty acids to the brain in LPC form. Overexpression is useful for studying MFSD2A in heterologous expression contexts. Important consideration: MFSD2A is principally expressed at the blood-brain barrier endothelium and in placenta — HCT 116 is not the physiological context for canonical MFSD2A function. The EDITGENE MFSD2A Knockout in HCT 116 is most useful for biochemistry, transport assays, and structure-function studies. MFSD2A mutations cause autosomal recessive primary microcephaly 15 (MCPH15) due to impaired DHA delivery during brain development — disease variant rescue (e.g., S339L, S399L) enables genotype-function studies. Rescue with wild-type or transport-deficient MFSD2A is the standard specificity control. The model is also relevant for studying MFSD2A's role in restricting transcytosis at the BBB.

Primary applications: • LPC-DHA transport: ³H-LPC-DHA or BODIPY-LPC uptake assays to quantify MFSD2A-dependent sodium-coupled omega-3 LPC transport. • Microcephaly modeling: rescue with patient-derived MFSD2A mutations (e.g., S339L) for genotype-function studies of primary microcephaly 15. • BBB-relevant studies: in heterologous BBB-relevant contexts, characterization of MFSD2A's selective LPC transport across endothelial membranes. • Transcytosis regulation: MFSD2A restricts transcytosis at the BBB — in BBB models, transcytosis frequency analysis in the absence of MFSD2A. EDITGENE recommends this model for in vitro MFSD2A biochemistry and structure-function studies; physiological BBB-relevant MFSD2A research requires brain microvascular endothelial cell models.

Yes. MFSD2A rescue experiments require attention to MFS family architecture: • Construct design: use a codon-modified MFSD2A sequence with a small intracellular tag (FLAG, HA). MFSD2A has 12 transmembrane domains characteristic of MFS transporters — preserve topology. • Surface localization validation: confirm plasma membrane localization by cell surface biotinylation or staining before functional assays. • Transport-deficient rescue: substrate-binding pocket mutations affecting LPC-DHA binding abolish transport activity and serve as the standard specificity control. • Microcephaly mutation rescue: patient-derived MFSD2A mutations enable disease genotype-function studies. • Functional readout: rescue should restore LPC-DHA uptake measured by radiolabeled or fluorescent LPC-DHA uptake assays. HCT 116 transduces efficiently with lentivirus and supports stable rescue line generation.