Transgene expression in PF4-surv and GATA-1-surv MKs and GATA-1-surv erythrocytes. (A) RT-PCR of the survivin transgene on RNA isolated from fresh BM of PF4-surv mice confirms survivin transgene expression. A single line of transgenic mice produced an abundant spliced transcript PCR product of about 300 bp in comparison to an unspliced size of 1.125 kb. Survivin transgene primers: sense 5′-cagccaccactttctgatag-3′ and antisense 5′-aaactggacagacagagagccaag-3′; GAPDH loading control primers: sense 5′-tcaccatcttccaggag-3′ and antisense 5′-gcttcaccaccttcttg-3′. (B) β-galactosidase transgene activity (blue) was highly detected in MKs from PF4-surv BM but not in other BM cells nor in control mouse BM, demonstrating lineage specificity of expression. Staining was performed in suspension in fixed cells before cells were cytospun and viewed via light microscopy. (C) Western blotting of magnetic bead-purified PF4-surv and wild-type (wt) MKs. MK marker, CD41, and β-actin were used as loading controls. (D) Western blotting of magnetic bead–purified GATA-1-surv MKs and MoFlo-sorted Ter119 + erythrocytes. β-actin and HSC70 were used as loading controls. Band quantitation using ImageJ 1.34s software (National Institutes of Health, Bethesda, MD) indicated an increase in transgenic survivin level over control in the range of 2- to 3-fold in PF4-suv MKs and 5- to 6-fold in GATA-1-surv MKs.