CAJ | 학술논문

目的构建包装分泌型人内皮抑素（Endostatin）基因过表达慢病毒并表达。方法人工合成含人胰岛素信号肽和人Endostatin基因序列，利用pLVX-mCMV-ZsGreen载体质粒，构建慢病毒pLVX-hEndostatin-mCMV-ZsGreen质粒，通过293细胞包装制备慢病毒。利用重组慢病毒，转染肝干细胞，筛选胞外表达分泌型Endostatin的肝干细胞，观察其能否稳定表达。结果成功构建pLVX-hEndostatin-mCMV-ZsGreen，酶切鉴定及测序正确；该载体可通过293细胞包装慢病毒，该病毒携带人胰岛素信号肽和Endostatin基因序列，在肝干细胞中稳定表达分泌。ELISA检测Endostatin表达水平为18．58～34．20 ng/ml。结论分泌型人Endostatin基因过表达慢病毒成功制备并表达，可用于肿瘤血管生成相关实验。
목적구건포장분비형인내피억소（Endostatin）기인과표체만병독병표체。방법인공합성함인이도소신호태화인Endostatin기인서렬，이용pLVX-mCMV-ZsGreen재체질립，구건만병독pLVX-hEndostatin-mCMV-ZsGreen질립，통과293세포포장제비만병독。이용중조만병독，전염간간세포，사선포외표체분비형Endostatin적간간세포，관찰기능부은정표체。결과성공구건pLVX-hEndostatin-mCMV-ZsGreen，매절감정급측서정학；해재체가통과293세포포장만병독，해병독휴대인이도소신호태화Endostatin기인서렬，재간간세포중은정표체분비。ELISA검측Endostatin표체수평위18．58～34．20 ng/ml。결론분비형인Endostatin기인과표체만병독성공제비병표체，가용우종류혈관생성상관실험。
Objective To construct and express the lentivirus over-expressing human secreted endostatin gene .Methods The synthetic human insulin signal peptide and endostatin gene sequences were created .pLVX-mC-MV-ZsGreen vector was used to develop lentivirus vector pLVX-hEndostatin-mCMV-ZsGreen .Recombinant lentivirus was produced in HEK 293 packaging cell line .Liver stem cells as target cells were transfected by recombinant lenti-virus,which expressed extracellular secreted endostatin ,and were screened for observing the stability of the cells ’ ex-pression .Results pLVX-hEndostatin-mCMV-ZsGreen lentivirus vector was developed successfully , and was con-firmed by restriction enzyme digestion and DNA sequencing .The vector could pack lentivirus with 293 cell,the lenti-virus expressed stably in the liver stem cells with human insulin signal peptide and endostatin gene sequences .The endostatin expression ranged from 18.58ng/ml to 34.20 ng/ml detected by ELISA.Conclusion Lentivirus over-ex-pressing secreted endostatin gene can be successfully developed and expressed in target cells ,which can be used for the research of tumor angiogenesis .