人 MEG3基因质粒载体的构建及其对人胰腺癌SW1990细胞增殖的影响  

Construction and inhibitory effect of MEG3 expression plasmid vector on the proliferation of human pancreatic carcinoma SW1990 cells

作　　者：陈瑞东[1] 唐文[1] 胡端敏[1] 

机构地区：[1]苏州大学附属第二医院消化内科,215004

出　　处：《国际肿瘤学杂志》2015年第7期488-491,共4页Journal of International Oncology

基　　金：苏州市科技计划（SYS201344）

摘　　要：目的：构建人类母源性印记基因3（MEG3）的真核表达载体，转染得到过表达的人胰腺癌SW1990细胞株，分析 MEG3过表达对 SW1990细胞增殖的影响。方法根据 GenBank 中 MEG3的基因序列，通过化学合成的方法进行全基因合成得到人 MEG3基因全长，将其构建入 pcDNA3.0真核表达载体中，得到 pcDNA3.0-MEG3表达载体，转染胰腺癌细胞 SW1990。采用 RT-PCR 及荧光定量 PCR 技术检测转染细胞中 MEG3基因的表达量。采用 MTT 试剂盒对转染后 SW1990细胞增殖能力的变化进行检测。实验中设转染 pcDNA3.0的 SW1990细胞为阴性对照组，普通 SW1990细胞为空白对照组。结果成功构建了 MEG3真核表达质粒pcDNA3.0-MEG3，并成功转染 SW1990细胞。与两对照组相比，转染后细胞的 MEG3基因表达量显著提高，提高约895倍（F ＝73.592，P ﹤0.01）。MTT 检测结果显示，MEG3基因明显抑制 SW1990细胞的增殖，实验组72 h 时吸光度值为0.81±0.06，与阴性对照组（1.17±0.07）和空白对照组（1.08±0.03）相比，3组差异有统计学意义（ F ＝33.489，P ﹤0.01）。结论本研究成功构建了MEG3真核表达质粒 pcDNA3.0-MEG3，并证实 MEG3基因及其产物对胰腺癌 SW1990细胞增殖有明显的抑制作用。Objective To construct a maternally expressed gene 3（MEG3）expression plasmid vector,and to obtain MEG3 over-expressed human pancreatic carcinoma SW1990 cells by transfection,and to analyze the effect of MEG3 overexpression on the proliferation of human pancreatic carcinoma SW1990 cells. Methods A complete gene sequence based on the sequence of MEG3 in the GenBank was designed and inserted into the eukaryotic expression vector pcDNA3. 0 to construct recombinant plasmid pcDNA3. 0-MEG3. It was identified by sequencing and transfected into human pancreatic carcinoma SW1990 cells. The expression of MEG3 in SW1990 cells was confirmed by RT-PCR. The effect of MEG3 on proliferation was evaluated by MTT assay. In this study,the SW1990 cells transfected by plasmid pcDNA3. 0 were named negative control group, and the usual SW1990 cells were named blank control group. Results A MEG3 expression plasmid vectorpcDNA3. 0-MEG3 was constructed successfully. And pcDNA3. 0-MEG3 vector was transfected into SW1990 cells successfully. The expression of MEG3 at mRNA in MEG3-SW1990 cells increased significantly,about 895 times（F = 73. 592,P ﹤ 0. 01）. The results of MTT assay indicated that over-expressed MEG3 could obviously inhibit SW1990 cells proliferation in vitro. After SW1990 cells transfected with pcDNA3. 0-MEG3 for 72 hours, the absorbance value was 0. 81 ± 0. 06,with a statistically significance（F = 33. 489,P ﹤ 0. 01）compared with negative control group（1. 17 ± 0. 07）and blank control group（1. 08 ± 0. 03）. Conclusion A MEG3 expression plasmid vector-pcDNA3. 0-MEG3 is constructed successfully. It is confirmed that MEG3 and its product have obvious inhibitory effects for the proliferation of human pancreatic carcinoma SW1990 cells.

关 键 词：胰腺肿瘤 RNA 基因表达调控 母源性印记基因3 

分 类 号：R735.9[医药卫生—肿瘤]