Semnan University of Medical Sciences

Research and Technology Vice Chancellor

Kues, Wilfried A. and Ahmadi, Samira and Davami, Fatemeh and Davoudi, Noushin and Nematpour, Fatemeh and Ahmadi, Maryam and Ebadat, Saeedeh and Azadmanesh, Kayhan and Barkhordari, Farzaneh and Mahboudi, Fereidoun (2017) Monoclonal antibodies expression improvement in CHO cells by PiggyBac transposition regarding vectors ratios and design. PLOS ONE, 12 (6). e0179902. ISSN 1932-6203

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Abstract

Establishing stable Chinese Hamster Ovary (CHO) cells producing monoclonal antibodies (mAbs) usually pass through the random integration of vectors to the cell genome, which is sensitive to gene silencing. One approach to overcome this issue is to target a highly transcribed region in the genome. Transposons are useful devices to target active parts of genomes, and PiggyBac (PB) transposon can be considered as a good option. In the present study, three PB transposon donor vectors containing both heavy and light chains were constructed, one contained independent expression cassettes while the others utilized either an Internal Ribosome Entry Site (IRES) or 2A element to express mAb. Conventional cell pools were created by transferring donor vectors into the CHO cells, whereas transposon-based cells were generated by transfecting the cells with donor vectors with a companion of a transposase-encoding helper vector, with 1:2.5 helper/donor vectors ratio. To evaluate the influence of helper/donor vectors ratio on expression, the second transposon-based cell pools were generated with 1:5 helper/donor ratio. Expression levels in the transposon-based cells were two to five -folds more than those created by conventional method except for the IRES-mediated ones, in which the observed difference increased more than 100-fold. The results were dependent on both donor vector design and vectors ratios.

Item Type: Article
Subjects: R Medicine > R Medicine (General)
Divisions: Faculty of Medical Sciences > School of Medicine
Depositing User: Mr Vahab Moshtaghi
Date Deposited: 28 Apr 2018 14:01
Last Modified: 21 May 2018 12:35
URI: http://eprints.semums.ac.ir/id/eprint/1281

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