慢性髓系白血病bcrabl融合基因在真核细胞中的表达  

THE EXPRESSION OF CHRONIC MYELOGENOUS LEUKEMIA bcr abl FUSION GENE IN EUKARYOTE

作　　者：刘楠[1] 孙秉中[1] 冯琦[1] 高杰英[2] 彭虹[2] 罗振 

机构地区：[1]第四军医大学西京医院,西安710032 [2]革军事医学科学院微生物流行病研究所

出　　处：《解放军医学杂志》2001年第6期426-428,共3页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金!(编号 39770 336 );陕西省科研基金!(编号98SM40 )

摘　　要：克隆慢性髓系白血病 (CML)融合基因bcr abl融合位点周围的cDNA序列 ,构建重组真核表达载体并表达于 p815细胞 ,探索bcr abl基因免疫治疗CML的可行性。设计两条引物扩增bcr abl融合位点周围 46 8bp的cDNA ,测序正确后 ,将其克隆至真核表达载体pcDNA 3.1,通过电穿孔法转染至 p815细胞中 ,应用G418筛选阳性细胞克隆 ,在含 10 %FCS的RPMI16 40中常规培养 ,收集细胞进行RT PCR鉴定。结果PCR扩增出了可用于基因免疫的位于bcr abl融合基因位点周围的 46 8bp的cDNA ,序列测定除第 44 8位碱基C突变为T外其余序列均正确 ;构建了重组真核表达载体 ,并用电穿孔法将其转染 p815细胞 ,RT PCR法检测到该细胞系有bcrChronic myelogenous leukemia bcr abl fusion gene was cloned and a recombinant expression vector was constructed and expressed in p815 cells to explore the feasibility of bcr abl fusion gene in CML gene immunization therapy. Two primers were designed for the amplification of a 468bp bcr abl cDNA coding sequence by PCR. After the PCR product was confirmed by the sequence analysis, it was properly inserted into the expression vextor pcDNA 3.1. The recombinant vector was transfected in p815 cells by means of electric perforation. These stable expression cell colonies were obtained by G418 selection ,and its expression was confirmed by RT PCR after cultivation in RPMI 1640. A 468bp bcr abl cDNA was amplified by PCR, and sequence analysis showed the whole correct sequence except base C mutated for T in position 452. A recombinant expression vector was constructed, and it could be tranfected into p815 cells by electric perforation. RT PCR examination showed that bcr abl had been transcribed in cell clones.

关 键 词：克隆 融合蛋白质类 BCR-ABL融合基因 慢性髓系白血病 真核细胞 肿瘤转移 

分 类 号：R733.72[医药卫生—肿瘤] Q786[医药卫生—临床医学]