论文部分内容阅读
目的:建立OPCML真核细胞表达质粒。方法:利用计算机软件Vector NTI,根据GenBank中OPCML的序列,用RT-PCR法从小鼠大脑组织中扩增OPCML基因全长片段,将其克隆入真核细胞表达质粒pcDNA3,建立OPCML的表达质粒pcDNA3-OPCML,后将其转染入人胚胎肾细胞(293T),用Western blot检测OPCML蛋白在293T中的表达。结果:OPCML能被成功地从正常组织中克隆,构建了真核细胞表达质粒pcDNA3-OPCML,测序表明其携带有正常OPCML的全长序列;转导入人类细胞后有OPCML蛋白的高量表达。结论:pcDNA3-OPCML携带有正常的OPCML全长基因序列,其能在真核细胞中高量表达功能蛋白。
Objective: To establish OPCML eukaryotic expression plasmid. Methods: According to the sequence of OPCML in GenBank, a full-length fragment of OPCML gene was amplified from mouse brain by RT-PCR and cloned into eukaryotic expression plasmid pcDNA3. The expression vector pcDNA3 -OPCML, and then transfected into human embryonic kidney cells (293T). Western blot was used to detect the expression of OPCML protein in 293T cells. Results: OPCML was cloned successfully from normal tissues. The eukaryotic expression plasmid pcDNA3-OPCML was constructed and sequenced. It showed that it carried the full-length sequence of normal OPCML. After transfected into human cells, OPCML protein was highly expressed. Conclusion: pcDNA3-OPCML carries the normal OPCML full length gene sequence, which can express high level of functional protein in eukaryotic cells.