Transfection, Spinfection, Exofection, and Luciferase Assays for Analysis of CCN Genes Expression Mechanism

Takanori Eguchi, Yanyin Lu, Eman A. Taha, Yuka Okusya

Research output: Chapter in Book/Report/Conference proceedingChapter

2 Citations (Scopus)

Abstract

Cell communication network factor 2 (CCN2), also known as connective tissue growth factor (CTGF), is protein inducible in response to TGFβ/Smad signal or the transcriptional activity of matrix metalloproteinase 3 (MMP3). We discovered that MMP3 in exosomes is transferable to recipient cells and then translocates into cell nuclei to transactivate the CCN2/CTGF gene. Exosomes and liposomes enable molecular transfection to recipient cells in vitro and in vivo. These small vesicles are surrounded by lipid membranes and carry proteins, RNA, DNA, and small chemicals. Here we define the exosome-based transfection as “exofection.” In addition, spinfection increases the efficiencies of transfection, exofection, and viral infection, thus being compatible with various molecular transfer protocols. Here, we provide protocols, tips, and practical examples of transfection, spinfection, exofection, fluorescence microscopy, and luciferase assays to analyze the CCNs gene expression mechanisms.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages103-126
Number of pages24
DOIs
Publication statusPublished - 2023

Publication series

NameMethods in Molecular Biology
Volume2582
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • cis-elements
  • enhancer
  • exofection
  • Exosomes
  • gaussia luciferase
  • gene promoter activity
  • immunofluorescence
  • luciferase assay
  • spinfection
  • transfection

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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