Seprate The Target Genes Of Cbfal With The PCR-selected Subtractive Hybridization

The development of the skeleton is a complicated process involving many factors. Transcription factors play a vital role in the developing process. To date, Cbfa1 (Core Binding Factor alpha 1) and Osx(osterix) have been identified as transcription factors which specifically express in the osteoblast lineage cells. In 1997, Cbfa1 which can activate the earlier differentiation stage of the osteoblast was proved to be a key factor for bone formation.Cbfa1 is not only a key regulator controlling bone formation and growth, but also important in cartilage maturation. And it may also be involved in osteoclastogenesis and cartilage vascular invasion. Moderate expression of Cbfa1 can promote the differentiation of the osteoblasts, but overexpression of Cbfa1 can suppress the mutation of the osteoblasts. It was in 2002 that Osx was identified by Kazuhisa Nakashima as another transcription factor by Suppression Subtractive Hybridization (SSH). Cbfa1 and Osx act at different stage during the differentiation of the osteoblasts. Cbfa1 has an essential role in the stage that Multipotential mesenchymal progenitors differentiate into preosteoblasts, and Osx is required in preosteoblasts differentiating into functional osteoblasts. Cbfa1 and Osx regulate the differentiation of osteoblast in different period. Cbfa1 can promote the differentiation of mesenchymal cells, and it can also induce the contra-differentiation of the differential cells. But Osx paly a important role in pre-osteoblast. The direct purpose of this article is to separate some new target genes induced by Cbfa1 in mesenchymal cells and osteocyte, and compare the similarities and differences of the genes. And its long-term aim is to identified another transcription factor. We transfected Cbfa1 into C3H10T1/2 a kind of mesenchymal cell and MLO-Y4 (Murine Long Bone Osteocyte-Y4) which has the characters of osteocyte. Then separate induced genes from them by SSH. We have constructed a library containing genes induced by Cbfa1 successfully. After partial sequencing of the individuals randomly in the library we get the sequences of the genes. Then we analyse and compare the result of sequence. We test the correctness and feasibility by Virtual Northern blot.