白血病·淋巴瘤
白血病·淋巴瘤
백혈병·림파류
JOURNAL OF LEUKEMIA & LYMPHOMA
2010年
2期
84-87
,共4页
唐友红%钟美佐%刘巍%黄进%李斌
唐友紅%鐘美佐%劉巍%黃進%李斌
당우홍%종미좌%류외%황진%리빈
启动区(遗传子)%转录启动子%基因载体%荧光素酶%淋巴瘤
啟動區(遺傳子)%轉錄啟動子%基因載體%熒光素酶%淋巴瘤
계동구(유전자)%전록계동자%기인재체%형광소매%림파류
Promoter regions(genetics)%Transcription initiation site%Gene vector%Luciferase%Lymphoma
目的 克隆人类生存蛋白(Survivin)核心启动子,研究Survivin启动子在人类淋巴瘤细胞Ramos和健康人肝脏细胞Chang Liver中的转录活性.方法 以人类肠基因组DNA为模板,PCR扩增Survivin启动子987 bp片段,将其通过酶切位点连入pGL3-Basic载体构建pGL3-Survivin荧光素酶报告基因载体,通过脂质体转染法转染Ramos和Chang Liver细胞,通过检测荧光素酶表达水平,比较Survivin启动子在这两种细胞中的转录活性.结果 成功克隆出987 bp的Survivin启动子;双酶切、PCR检测和DNA测序证实pGL3-Survivin载体构建成功;荧光素酶活性检查显示:Survivin启动子在Ramos细胞中的转录活性为阳性对照CMV启动子活性的4.5%,明显高于在Chang Liver细胞中的0.19%,在淋巴瘤细胞中具有较高特异性,且在淋巴瘤细胞中的活性明显高于阴性对照pGL3-Basic的0.086%.结论 Survivin启动子在淋巴瘤细胞中具有较高的特异性,可以作为淋巴瘤细胞基因转染的肿瘤特异性启动子使用.
目的 剋隆人類生存蛋白(Survivin)覈心啟動子,研究Survivin啟動子在人類淋巴瘤細胞Ramos和健康人肝髒細胞Chang Liver中的轉錄活性.方法 以人類腸基因組DNA為模闆,PCR擴增Survivin啟動子987 bp片段,將其通過酶切位點連入pGL3-Basic載體構建pGL3-Survivin熒光素酶報告基因載體,通過脂質體轉染法轉染Ramos和Chang Liver細胞,通過檢測熒光素酶錶達水平,比較Survivin啟動子在這兩種細胞中的轉錄活性.結果 成功剋隆齣987 bp的Survivin啟動子;雙酶切、PCR檢測和DNA測序證實pGL3-Survivin載體構建成功;熒光素酶活性檢查顯示:Survivin啟動子在Ramos細胞中的轉錄活性為暘性對照CMV啟動子活性的4.5%,明顯高于在Chang Liver細胞中的0.19%,在淋巴瘤細胞中具有較高特異性,且在淋巴瘤細胞中的活性明顯高于陰性對照pGL3-Basic的0.086%.結論 Survivin啟動子在淋巴瘤細胞中具有較高的特異性,可以作為淋巴瘤細胞基因轉染的腫瘤特異性啟動子使用.
목적 극륭인류생존단백(Survivin)핵심계동자,연구Survivin계동자재인류림파류세포Ramos화건강인간장세포Chang Liver중적전록활성.방법 이인류장기인조DNA위모판,PCR확증Survivin계동자987 bp편단,장기통과매절위점련입pGL3-Basic재체구건pGL3-Survivin형광소매보고기인재체,통과지질체전염법전염Ramos화Chang Liver세포,통과검측형광소매표체수평,비교Survivin계동자재저량충세포중적전록활성.결과 성공극륭출987 bp적Survivin계동자;쌍매절、PCR검측화DNA측서증실pGL3-Survivin재체구건성공;형광소매활성검사현시:Survivin계동자재Ramos세포중적전록활성위양성대조CMV계동자활성적4.5%,명현고우재Chang Liver세포중적0.19%,재림파류세포중구유교고특이성,차재림파류세포중적활성명현고우음성대조pGL3-Basic적0.086%.결론 Survivin계동자재림파류세포중구유교고적특이성,가이작위림파류세포기인전염적종류특이성계동자사용.
Objective To clone core promoter of human Survivin gene,and investigate the transcriptional activity of the Survivin promoter in the human lymphoma cells Ramos and the normal human liver cells Chang Liver.Methods The Survivin promoter was amplified by PCR using the total genomic DNA of human colon as sample DNA and then cloned into pGL3-Basic vector.The reporter gene vector was named pGL3-Survivin.This vector was transfected into Ramos cells and the control Chang Liver cells to detect the transcriptional activities of Survivn promoter by cationic liposome.Results The Survivin promoter was amplified through PCR successfully.The results of restriction enzyme digestion,PCR and sequencing indicated that the pGL3-Survivin luciferase expression vector was constructed triumphantly.Survivin promoter showed much more higher transcriptional activity in Ramos cells than in Chang Liver cells.Relatively,pGL3CMV and pGL3-Basic separately showed no obviously different transcriptional activity in Ramos cells and Chang Liver cells.Conslusion The Survivn promoter has lymphoma specificity and may be used as tumor specific promoter in gene transfection of lymphoma.
