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SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells

Chinese hamster ovary (CHO) cells have become the most widely utilized mammalian cell line for the production of recombinant proteins. However, the product yield and transgene instability need to be further increased and solved. In this study, we investigated the effect of five different introns on...

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Autores principales: Xu, Dan‐hua, Wang, Xiao‐yin, Jia, Yan‐long, Wang, Tian‐yun, Tian, Zheng‐wei, Feng, Xin, Zhang, Yin‐na
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5867124/
https://www.ncbi.nlm.nih.gov/pubmed/29441681
http://dx.doi.org/10.1111/jcmm.13504
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author Xu, Dan‐hua
Wang, Xiao‐yin
Jia, Yan‐long
Wang, Tian‐yun
Tian, Zheng‐wei
Feng, Xin
Zhang, Yin‐na
author_facet Xu, Dan‐hua
Wang, Xiao‐yin
Jia, Yan‐long
Wang, Tian‐yun
Tian, Zheng‐wei
Feng, Xin
Zhang, Yin‐na
author_sort Xu, Dan‐hua
collection PubMed
description Chinese hamster ovary (CHO) cells have become the most widely utilized mammalian cell line for the production of recombinant proteins. However, the product yield and transgene instability need to be further increased and solved. In this study, we investigated the effect of five different introns on transgene expression in CHO cells. hCMV intron A, adenovirus tripartite leader sequence intron, SV40 intron, Chinese hamster EF‐1alpha gene intron 1 and intervening sequence intron were cloned downstream of the eGFP expression cassette in a eukaryotic vector, which was then transfected into CHO cells. qRT‐PCR and flow cytometry were used to explore eGFP expression levels. And gene copy number was also detected by qPCR, respectively. Furthermore, the erythropoietin (EPO) protein was used to test the selected more strong intron. The results showed that SV40 intron exhibited the highest transgene expression level among the five compared intron elements under transient and stable transfections. In addition, the SV40 intron element can increase the ratio of positive colonies and decrease the coefficient of variation in transgene expression level. Moreover, the transgene expression level was not related to the gene copy number in stable transfected CHO cells. Also, the SV40 intron induced higher level of EPO expression than IVS intron in transfected CHO cell. In conclusion, SV40 intron is a potent strong intron element that increases transgene expression, which can readily be used to more efficient transgenic protein production in CHO cells.
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spelling pubmed-58671242018-04-01 SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells Xu, Dan‐hua Wang, Xiao‐yin Jia, Yan‐long Wang, Tian‐yun Tian, Zheng‐wei Feng, Xin Zhang, Yin‐na J Cell Mol Med Original Articles Chinese hamster ovary (CHO) cells have become the most widely utilized mammalian cell line for the production of recombinant proteins. However, the product yield and transgene instability need to be further increased and solved. In this study, we investigated the effect of five different introns on transgene expression in CHO cells. hCMV intron A, adenovirus tripartite leader sequence intron, SV40 intron, Chinese hamster EF‐1alpha gene intron 1 and intervening sequence intron were cloned downstream of the eGFP expression cassette in a eukaryotic vector, which was then transfected into CHO cells. qRT‐PCR and flow cytometry were used to explore eGFP expression levels. And gene copy number was also detected by qPCR, respectively. Furthermore, the erythropoietin (EPO) protein was used to test the selected more strong intron. The results showed that SV40 intron exhibited the highest transgene expression level among the five compared intron elements under transient and stable transfections. In addition, the SV40 intron element can increase the ratio of positive colonies and decrease the coefficient of variation in transgene expression level. Moreover, the transgene expression level was not related to the gene copy number in stable transfected CHO cells. Also, the SV40 intron induced higher level of EPO expression than IVS intron in transfected CHO cell. In conclusion, SV40 intron is a potent strong intron element that increases transgene expression, which can readily be used to more efficient transgenic protein production in CHO cells. John Wiley and Sons Inc. 2018-02-14 2018-04 /pmc/articles/PMC5867124/ /pubmed/29441681 http://dx.doi.org/10.1111/jcmm.13504 Text en © 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Articles
Xu, Dan‐hua
Wang, Xiao‐yin
Jia, Yan‐long
Wang, Tian‐yun
Tian, Zheng‐wei
Feng, Xin
Zhang, Yin‐na
SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title_full SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title_fullStr SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title_full_unstemmed SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title_short SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells
title_sort sv40 intron, a potent strong intron element that effectively increases transgene expression in transfected chinese hamster ovary cells
topic Original Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5867124/
https://www.ncbi.nlm.nih.gov/pubmed/29441681
http://dx.doi.org/10.1111/jcmm.13504
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