STK38L Knockout HAP1 Cell Line

The choice depends on whether you are studying STK38L's role as a Hippo pathway-related NDR kinase or its functions in centrosome regulation, cell cycle, and immune cell biology. The Knockout line is appropriate for asking whether STK38L is required for substrate phosphorylation in contexts where its STK38 (NDR1) paralog may not fully compensate. Overexpression is useful for testing whether elevated STK38L activity drives specific phenotypes or for studying its hippo-related signaling contributions. Important consideration: STK38 (NDR1) and STK38L (NDR2) share substantial functional overlap as NDR family kinases. Single STK38L knockout in HAP1 may show mild phenotypes if STK38 compensates. The EDITGENE Knockout is most informative when combined with the parallel STK38 knockout (also available) for family-specific dissection. Rescue with wild-type or kinase-dead STK38L is the standard control.

Primary applications: • NDR substrate phosphorylation: phospho-substrate analysis (FoxO, c-Myc, MOB1) to assess STK38L-dependent phosphorylation events. • Centrosome regulation: imaging-based analysis of centrosome number, duplication, and separation. • Cell cycle progression: flow cytometry-based cell cycle analysis and mitotic phenotyping. • Paralog studies: combined analysis with the parallel STK38 knockout (also available) dissects NDR1/NDR2 family-specific functions. EDITGENE recommends this model for researchers investigating NDR kinase biology, Hippo pathway-related signaling, and centrosome regulation.

Yes. STK38L rescue experiments require attention to NDR activation requirements: • Construct design: use a codon-modified STK38L sequence with a small C-terminal tag (FLAG, HA). The N-terminal regulatory region (NTR) interacts with MOB1 activators and must be preserved. • Kinase-dead rescue: an active site mutation (typically K117A or D203A in the catalytic loop) abolishes catalytic activity and is the standard specificity control. • MOB1-binding-deficient rescue: NTR mutations that disrupt MOB1 binding test the dependence of phenotypes on NDR kinase activation rather than basal activity. • Paralog rescue panel: rescue with STK38L in STK38 knockouts (or combined-knockout backgrounds) enables family-specific function dissection. HAP1-specific considerations: • Diploidization: HAP1 cells gradually diploidize during extended culture — confirm ploidy by flow cytometry at the time of phenotypic assay. • Integration site sensitivity: position effects on transgene expression are more pronounced in near-haploid backgrounds; generating multiple independent rescue clones is strongly recommended. • Transduction efficiency: HAP1 transduces with lentivirus at moderate efficiency — increase MOI compared to standard immortalized lines.