医学分子生物学杂志2025，Vol.22Issue(5)：415-422,8.DOI:10.3870/j.issn.1672-8009.2025.05.001
BMP-2生物学活性新型检测系统的构建及功能验证
Construction and Functional Validation of a Novel Biological Activity Detection System for BMP-2
摘要
Abstract
Objective To develope a novel BMP-2 bioactivity detection system based on a du-al-luciferase reporter gene stably transfected cell line(C2C12BBDE)to address the issues of low sen-sitivity,time-consuming procedures in traditional alkaline phosphatase(ALP)activity assays,and cumbersome operational workflows in real-time fluorescent quantitative polymerase chain reaction(qRT-PCR)methods.Methods This system integrates a BMP-2 responsive promoter driving NanoLuc luciferase(NLuc)expression with constitutively expressed Firefly luciferase(Fluc)as an internal control via lentiviral transduction.Potential promoter interference was mitigated through strategic design of an inverse transcription unit and Poly A sequence insertion.Results This novel system achieved a dynamic detection range of 0-30 ng/mL,matching the sensitivity of qRT-PCR while demonstrating a statistically significant improvement over the ALP assay.Furthermore,the as-say cycle was markedly reduced to 16 hours,representing an efficiency enhancement exceeding 700％relative to the ALP method.Critically,the workflow is streamlined,obviating the need for complex RNA extraction and reverse transcription steps inherent in qRT-PCR.Conclusion The successful establishment of this system not only furnishes a standardized,high-throughput tool for BMP-2 bioactivity assessment but also provides a modular,adaptable technical paradigm for the de-velopment of bioactivity assays for other growth factors.This advancement holds substantial transla-tional potential,significantly contributing to the rational development and clinical implementation of bone regenerative therapeutics.关键词
骨形态发生蛋白-2/双萤光素酶报告系统/纳米萤光素酶/生物活性检测Key words
bone morphogenetic protein-2/dual-luciferase reporter assay/nanoluciferase/bioactivity detection分类
生物科学引用本文复制引用
符岳阳,夏芷毅,杨建勋,王碧茹,郑瑞,阎博..BMP-2生物学活性新型检测系统的构建及功能验证[J].医学分子生物学杂志,2025,22(5):415-422,8.基金项目
国家肿瘤生物学重点实验室重点课题(No.CBSKL2022ZZ04),国家肿瘤生物学重点实验室自主课题(No.2025GTEP012),陕西省重点项目-实验室重点项目(No.2025SYS-SYSZD-030) This work was supported by grants from the National Tumor Biology Key Laboratory(No.CBSKL2022ZZ04,No.2025GTEP012),Key Laboratory Pro-gram from Shaanxi Province(No.2025SYS-SYSZD-030) （No.CBSKL2022ZZ04）
