山东医药
山東醫藥
산동의약
SHANDONG MEDICAL JOURNAL
2014年
9期
20-22
,共3页
柳君如%张峻岭%徐士福%程琳
柳君如%張峻嶺%徐士福%程琳
류군여%장준령%서사복%정림
脂质体%DMRIE-C%小干扰RNA%Jurkat细胞
脂質體%DMRIE-C%小榦擾RNA%Jurkat細胞
지질체%DMRIE-C%소간우RNA%Jurkat세포
liposomes%DMRIE-C%small interfering RNA%Jurkat cells
目的:探讨由DMRIE-C脂质体介导将CLEC2B siRNA转染入Jrukat细胞的过程中,对Jurkat细胞的影响,并分析其转染率的影响因素。方法通过DMRIE-C转染CLEC2B siRNA入Jrukat细胞,台盼蓝染色法观察细胞活性,流式细胞仪检测siRNA转染率,RT-PCR检测CLEC2B基因沉默效率。结果 DMRIE-C脂质体对细胞的损伤程度与两者比例相关,对转染体系600μL (其中200μL密度为2×106/mL淋巴细胞)进行转染时,脂质体达3μL时可损伤细胞,不利于后续试验进行;siRNA∶DMRIE-C比例为3μL∶2μL,siRNA终浓度约80 nmol/L时,转染率最高,可达30.7%,转染后CLEC2B基因抑制率可达(35±1.9)%(t=15.9,P<0.05)。结论 DMRIE-C脂质体可实现目的siRNA对Jurkat细胞有效转染,同时转染中脂质体与细胞比例、血清、操作方式等因素均可对转染效率产生一定的影响。
目的:探討由DMRIE-C脂質體介導將CLEC2B siRNA轉染入Jrukat細胞的過程中,對Jurkat細胞的影響,併分析其轉染率的影響因素。方法通過DMRIE-C轉染CLEC2B siRNA入Jrukat細胞,檯盼藍染色法觀察細胞活性,流式細胞儀檢測siRNA轉染率,RT-PCR檢測CLEC2B基因沉默效率。結果 DMRIE-C脂質體對細胞的損傷程度與兩者比例相關,對轉染體繫600μL (其中200μL密度為2×106/mL淋巴細胞)進行轉染時,脂質體達3μL時可損傷細胞,不利于後續試驗進行;siRNA∶DMRIE-C比例為3μL∶2μL,siRNA終濃度約80 nmol/L時,轉染率最高,可達30.7%,轉染後CLEC2B基因抑製率可達(35±1.9)%(t=15.9,P<0.05)。結論 DMRIE-C脂質體可實現目的siRNA對Jurkat細胞有效轉染,同時轉染中脂質體與細胞比例、血清、操作方式等因素均可對轉染效率產生一定的影響。
목적:탐토유DMRIE-C지질체개도장CLEC2B siRNA전염입Jrukat세포적과정중,대Jurkat세포적영향,병분석기전염솔적영향인소。방법통과DMRIE-C전염CLEC2B siRNA입Jrukat세포,태반람염색법관찰세포활성,류식세포의검측siRNA전염솔,RT-PCR검측CLEC2B기인침묵효솔。결과 DMRIE-C지질체대세포적손상정도여량자비례상관,대전염체계600μL (기중200μL밀도위2×106/mL림파세포)진행전염시,지질체체3μL시가손상세포,불리우후속시험진행;siRNA∶DMRIE-C비례위3μL∶2μL,siRNA종농도약80 nmol/L시,전염솔최고,가체30.7%,전염후CLEC2B기인억제솔가체(35±1.9)%(t=15.9,P<0.05)。결론 DMRIE-C지질체가실현목적siRNA대Jurkat세포유효전염,동시전염중지질체여세포비례、혈청、조작방식등인소균가대전염효솔산생일정적영향。
Objective To observe the process of transfecting CLEC 2B siRNA mediated by DMRIE-C liposome into the Jurkat cells and to analyze the factors of influencing transfection rate .Methods We transfected CLEC2B siRNA into the Jurkat cells by DMRIE-C.Trypan blue staining was used to observe the cell activity .siRNA transfection efficiency was de-tected by flow cytometry , and the CLEC2B gene silencing efficiency was detected by RT-PCR.Results The cell damage degree in the transfection process was influenced by the cell members /transfection system lipo concentration ratio , 3μl lipo in the transfection system (600 μl total liquid, 200 μl 2 ×106/ml lymphocytes) would damage cells, and go against the following experiment.The transfection efficiency could increase to 30.70%when the siRNA:DMRIE-C ratio was adjusted to 3 μl∶2 μl, and then, the CLEC2B gene expression inhibition ratio was about 35 ±1.9%(t=15.9, P<0.05).Con-clusion DMRIE-C lipo cloud successfully transfer aim siRNA to Jurkat cells , and meanwhile, lipo/lymphocyte ratio, ser-um interference and operation skills may influence the transfection efficiency .