CXCL12 Stimulation of CXCR4-WHIM Transduced Waldenstrom's Macroglobulinemia cell line BCWM.1

CXCR4 cDNA transcripts were subcloned into plenti-IRES-GFP vectors, and stably transduced using a lentiviral system In addition to wild-type CXCR4, vectors with the mutations c.1013C>G (p.Ser338*), c.932_933insT (p.Thr311fs), and c.1030_1041delinsGT (p.Ser344fs) were generated. Cell lines were stimulated with 50nM of the CXCR4 ligand CXCL12 (SDF1A) (R&D Systems, Minneapolis MN) for two hours. RNA from each sample was extracted at baseline and at 2 hours. At baseline and following stimulation, RNA was extracted and processed for analysis using Affymetrix Human Gene ST 2.0 arrays (Affymetrix, Santa Clara, CA). Data was analyzed using Oligo and Limma bioconductor packages in R using TMM normalization.