Current Topics in TransplantationExperimental studiesIdentification of Suitable Reference Genes for mRNA Studies in Bone Marrow in a Mouse Model of Hematopoietic Stem Cell Transplantation

- During HSCT in bone marrow, ACTB displayed the highest expression in the combined group (all samples).
- During HSCT in bone marrow, the lowest expression of PSDHA was found in the TBI group.
- During HSCT in bone marrow, the lowest expression of HPRT was found in the BU/CY group and the combined group.
- GeNorma and NormFinder showed B2M as the most stably expressed gene.
- B2M was recommended as the most suitable reference gene after comprehensive ranking.

BackgroundBone marrow micro-environment changes during hematopoietic stem cell transplantation (HSCT) with subsequent alteration of genes expression. Quantitative polymerase chain reaction (q-PCR) is a reliable and reproducible technique for the analysis of gene expression. To obtain more accurate results, it is essential to find a reference during HSCT. However, which gene is suitable during HSCT remains unclear. This study aimed to identify suitable reference genes for mRNA studies in bone marrow after HSCT.MethodsC57BL/6 mice were treated with either total body irradiation (group T) or busulfan/cyclophosphamide (BU/CY) (group B) followed by infusion of bone marrow cells. Normal mice without treatments were served as a control. All samples (group T + group B + control) were defined as group G. On days 7, 14, and 21 after transplantation, transcription levels of 7 candidate genes, ACTB, B2M, GAPDH, HMBS, HPRT, SDHA, and YWHAZ, in bone marrow cells were measured by use of real-time quantitative PCR. The expression stability of these 7 candidate reference genes were analyzed by 2 statistical software programs, GeNorm and NormFinder.ResultsOur results showed that ACTB displayed the highest expression in group G, with lowest expression of PSDHA in group T and HPRT in groups B and G. Analysis of expression stability by use of GeNorm or NormFinder demonstrated that expression of B2M in bone marrow were much more stable during HSCT, compared with other candidate genes including commonly used reference genes GAPDH and ACTB.ConclusionsACTB could be used as a suitable reference gene for mRNA studies in bone marrow after HSCT.