一种利用瞬时转染进行入侵相关基因分析的新生物测定方法。

N Platet, M Garcia
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引用次数: 12

摘要

为了理解肿瘤侵袭和转移的机制,需要模型系统来分离这些复杂的、多方面的过程的各个步骤。我们提出了一种新的方法来鉴定参与细胞侵袭和/或运动的基因,该方法具有瞬时基因转染和Matrigel侵袭试验的综合优势。将表达目标基因和荧光素酶的两种载体瞬时共转染癌细胞,作为转染细胞的标记,然后检测Matrigel侵袭。荧光素酶共转染似乎是一种敏感的半定量检测转染细胞,并在整个入侵试验中是最大的。使用磷酸钙沉淀的转染程序不影响细胞侵袭性,并允许两种基因的细胞共表达。应用该方法,我们发现未配体和配体人雌激素受体α的瞬时表达可阻止MDA-MB-231乳腺癌细胞的侵袭性。总之,我们提出了快速和通用的体外程序来研究单个克隆基因对细胞过程的影响,如侵袭和运动。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
A new bioassay using transient transfection for invasion-related gene analysis.

To understand the mechanisms of tumor invasion and metastasis, model systems are required that isolate the individual steps of these complicated, multifaceted processes. We propose a new procedure to identify genes involved in cell invasion and/or motility that features the combined advantages of transient gene transfection and Matrigel invasion assays. Cancer cells were transiently cotransfected with two vectors expressing the gene of interest and luciferase, as a marker of transfected cells, and then assayed for Matrigel invasion. Luciferase cotransfection appeared to be a sensitive semiquantitative assay for transfected cells and was maximal throughout the invasion assay. The proposed transfection procedure, using calcium phosphate precipitation, did not affect cell invasiveness and allowed cellular coexpression of both genes. When applying this method, we found that transient expression of the unliganded and liganded human estrogen receptor alpha prevented invasiveness of MDA-MB-231 breast cancer cells. In conclusion, we propose rapid and versatile in vitro procedure for studying the effects of individual cloned genes on cellular processes, such as invasion and motility.

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