CAJ | 학술논문

目的通过克隆转录因子SPDEF以及其真核表达载体的构建,分析该基因对成釉细胞MMP-20基因表达的影响,为进一步研究转录因子SPDEF在牙釉质形成中发挥的作用奠定基础.方法利用RT-PCR技术从小鼠成釉细胞中获得SPDEF基因,测序正确后克隆至pcDNA3.1/myc-HisA载体中,将重组质粒瞬时转染成釉细胞,并用双荧光素酶报告基因系统检测技术来检测MMP-20启动子区域的转录活性.结果①SPDEF基因经过RT-PCR扩增得到的产物与预期片段978bp相符,将所获得的目的基因与GenBank 提供的已知序列(NM:013891.4)完全一致.②重组质粒pcDNA3.1/myc-HisA-SPDEF转染成釉细胞后,用荧光素酶分析系统检测显示MMP-20启动子转录活性升高.结论成功构建了SPDEF真核表达载体,初步研究证明SPDEF可能与MMP-20特征性序列结合从而上调MMP-20的表达.
목적통과극륭전록인자SPDEF이급기진핵표체재체적구건,분석해기인대성유세포MMP-20기인표체적영향,위진일보연구전록인자SPDEF재아유질형성중발휘적작용전정기출.방법이용RT-PCR기술종소서성유세포중획득SPDEF기인,측서정학후극륭지pcDNA3.1/myc-HisA재체중,장중조질립순시전염성유세포,병용쌍형광소매보고기인계통검측기술래검측MMP-20계동자구역적전록활성.결과①SPDEF기인경과RT-PCR확증득도적산물여예기편단978bp상부,장소획득적목적기인여GenBank 제공적이지서렬(NM:013891.4)완전일치.②중조질립pcDNA3.1/myc-HisA-SPDEF전염성유세포후,용형광소매분석계통검측현시MMP-20계동자전록활성승고.결론성공구건료SPDEF진핵표체재체,초보연구증명SPDEF가능여MMP-20특정성서렬결합종이상조MMP-20적표체.
Objective To analyse the influence of the expression of MMP-20 gene by cloning the transcription factor SPDEF gene and constructing eukaryotic expression vector in order to further study the role of transcription factor SPDEF in the formation of the en -amel.Methods The SPDEF gene was acquired from ALC by RT-PCR,Then inserted into pcDNA3.1/myc-HisA vector.The recombinant plasmid was transiently transfected into ALC .Dual-Luciferase Reporter Assay System was performed to detect the transcriptional activity of MMP-20 promoter region.Results ①SPDEF was amplification by RT-PCR and matched the expected fragment about 978bp.The sequencing result was completely matched with the known sequence in GeneBank accession NO .NM:013891.4.②The recombinant plasmid of pcDNA3. 1/myc-SPDEF was transfect into ALC.Then detect the increase of the transcriptional activity of MMP-20.Conclusion The eukaryotic ex-pression vector carried SPDEF is constructed successfully .Preliminary analysis has shown that SPDEF may be combined with the characteris-tic sequence of MMP-20 which up-regulate the expression of MMP-20.