抗山羊ΔfosB基因表达产物抗体的制备及应用

Preparation and application of goat ΔfosB gene expression product antibody

摘要:

ΔfosB是fosB基因的自然截短型，稳定存在于许多组织中，在脂肪细胞和成骨细胞的形成和分化中起到重要作用。ΔFosB蛋白可能与钙在骨和乳腺中的代谢有关，并且调控钙从骨向乳腺转移的信号通路。将奶山羊的ΔfosB基因亚克隆到pET32a载体得到pET32a-ΔfosB原核表达载体，IPTG诱导其在大肠杆菌中表达，纯化融合蛋白免疫兔子制备多克隆抗体。ELISA检测显示抗体效价达1∶51 200，Western blotting 结果显示制备的抗体能特异性检测原核表达的ΔFosB蛋白以及在HEK-293细胞中表达的ΔFosB蛋白。进一步的组织差异表达检测表明ΔFosB在山羊的乳腺、骨髓、脑髓、肌肉、心脏、肝和肺组织中均有表达。

Abstract:

ΔfosB, a naturally occurring truncated isform of fosB gene, existed in many tissues stably and played an important role in formation and differentiation of adipocyte and osteoblast. ΔFosB may be related to the metabolism of calcium in bone and mammary gland and regulate the signal pathway of calcium transfer from bone to mammary gland. We first sub-cloned ΔfosB gene of goat into the vector pET32a to construct prokaryotic expression vector pET32a-ΔfosB. Then we induced for ΔfosB gene expression efficiently by IPTG. Finally we immunized the adult rabbits with purified recombinant ΔFosB to prepare rabbit anti-goat ΔFosB polyclonal antibody. iELISA analysis showed the antibody with the titer of 1:51 200, and Western blotting result showed that the antibody could specifically detect the ΔFosB protein expressed in prokaryotic cell and HEK-293 cell, respectively. Further Western blotting assay showed that ΔFosB expressed in various tissues of goat in vivo.