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10. [Effect of Intron Orientation on the Expression of Transgene Imposed by MAR Expression Vector in Stably Recombinant CHO Cells].

Author

Li Q, Zhao CP, Wang XY, Sun QL, and Wang TY

Subjects
Animals, CHO Cells, Cricetinae, Cricetulus, Gene Dosage, Transfection, Genetic Engineering methods, Genetic Vectors, Introns, Matrix Attachment Regions, Transgenes
Abstract

Objective: To determine the effect of intron orientation on the transgene expression level imposed by matrix attachment region (MAR) expression vector., Methods: The MAR of β-globin was amplified by PCR, and then cloned into MAR expression vectors. An intron sequence was digested with restriction enzyme, ligated to the MAR expression vector in reverse orientation, and then transfected into Chinese hamster ovary (CHO) cells. The transfected stable cells were screened by G418. The level of chloramphenicol acetyltransferase (CAT) gene expression was analyzed by ELISA method., Results: The transgene expression levels of CHO cells with the two expression vectors with a positive intron or without MAR were higher than that of CHO cells with an expression vector with reverse intron (P < 0.05). MAR did not improve transgene expression with reverse intron presence., Conclusion: Different orientation of intron can affect transgene expression in recombinant CHO cells. The transgene expression level can be increased using positive intron and MAR.

Published
2016

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