Generation of recombinant adenoviruses

Nobuho Tanaka; Hirotaka Tsuno; Satoru Ohashi; Mitsuyasu Iwasawa; Hiroshi Furukawa; Tomohiro Kato; Naoshi Fukui

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Generation of recombinant adenoviruses

NT Nobuho Tanaka

HT Hirotaka Tsuno

SO Satoru Ohashi

MI Mitsuyasu Iwasawa

HF Hiroshi Furukawa

TK Tomohiro Kato

NF Naoshi Fukui

This method is extracted from research article: BMC Musculoskelet Disord, Feb 2021

The attenuation of insulin-like growth factor signaling may be responsible for relative reduction in matrix synthesis in degenerated areas of osteoarthritic cartilage

DOI: 10.1186/s12891-021-04096-w

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Recombinant adenoviruses were constructed using a ViraPower Adenoviral Expression System (Thermo Fisher Scientific). For this, human SP1 complementary DNA (cDNA) were cloned into the adenoviral-generating constructs [25]. The construct was then transfected into 293A cells (Thermo Fisher Scientific) using FuGENE 6 (Roche Diagnostics), and the cells were subcultured to generate recombinant adenoviruses carrying these genes under the control of the human cytomegalovirus immediate‐early enhancer/promoter. The viruses were titrated by limiting dilution plaque titration on 293A cells, and used at 50–100 plaque‐forming units/cell. Our preliminary experiments showed that the efficiency of transduction by this method is almost 100 %.

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