摘要
通过对体外传代培养的人血管起源平滑肌肉瘤细胞进行直接和脂质体Tfx-50包裹的人心钠素(ANF)基因表达质粒pcDNA3/ANF转染,并以非同位素地高辛标记的RNA分子探针对转染细胞中的ANFmRNA水平进行RNADotBlot分子杂交检测,同时利用放射免疫方法测定转染细胞培养上清中的ANF含量。结果显示,与未经脂质体Tfx-50包裹的pcDNA3/ANF直接进行细胞转染及低浓度脂质体Tfx-50包裹的pcDNA3/ANF进行细胞转染相比,脂质体Tfx-50与pcDNA3/ANF以3∶1(荷质比)进行包裹,并当pcDNA3/ANF在细胞数为105达到2.0μg时,细胞中ANFmRNA表达阳性,转染细胞培养上清中ANF含量达到58.0ng/L。表明通过脂质体Tfx-50介导可以提高pcDNA3/ANF的转染效率,并使细胞表达ANF显著增加。
Both Tfx 50 liposome coated and uncoated pcDNA3/ANF containing the human ANF gene were transfected into leiomyosarcoma (LMS) cells in vitro. The ANF mRNA of the LMS cells was detected by Dot Bloting method using the Diglabeled RNA probe. The ANF peptide levels in the LMS cell culture media were determined by RIA. The results showed that on day 3 after the gene transfection the ANF mRNA expression was positive and the ANF concentration of the cell culture media was up to 58 ng/L when pcDNA3/ANF DNA was coated with Tfx 50 under the charge ratio of 3∶1 (Tfx 50 liposome: pcDNA3/ANF DNA) and the quantity of the DNA was 2.0 μg/10 5 cell. These indicate that Tfx 50 reagent can be used to enhance pcDNA3/ANF transfection efficiency and gene expression, and it may be used for the study of ANF gene therapy on cardiovascular obstructive disease.
出处
《南京医科大学学报(自然科学版)》
CAS
CSCD
1999年第2期107-110,共4页
Journal of Nanjing Medical University(Natural Sciences)
基金
江苏省教委自然基金