Jun Cheng , Yanmin Zhang , Yuan Tian , Lei Cao , Xuping Liu , Shiwei Miao , Liang Zhao , Qian Ye , Yan Zhou , Wen-Song Tan
{"title":"Development of a novel tyrosine-based selection system for generation of recombinant Chinese hamster ovary cells","authors":"Jun Cheng , Yanmin Zhang , Yuan Tian , Lei Cao , Xuping Liu , Shiwei Miao , Liang Zhao , Qian Ye , Yan Zhou , Wen-Song Tan","doi":"10.1016/j.jbiosc.2023.12.013","DOIUrl":null,"url":null,"abstract":"<div><p><span><span><span>Efficiently expanding Chinese hamster ovary (CHO) cells, which serve as the primary host cells for </span>recombinant protein production, have gained increasing industrial significance. A significant hurdle in stable cell line development is the low efficiency of the target gene integrated into the host genome, implying the necessity for an effective screening and selection procedure to separate these stable cells. In this study, the genes of </span>phenylalanine hydroxylase<span><span> (PAH) and pterin 4 alpha carbinolamine dehydratase 1 (PCBD1), which are key </span>enzymes<span> in the tyrosine synthesis pathway, were utilized as selection markers and transduced into host cells together with the target genes. This research investigated the enrichment effect of this system and advanced further in understanding its benefits for cell line development and rCHO cell culture. A novel tyrosine-based selection system that only used </span></span></span><em>PCBD1</em><span> as a selection marker was designed to promote the enrichment effect. Post 9 days of starvation, positive transductants in the cell pool approached 100%. Applied the novel tyrosine-based selection system, rCHO cells expressing E2 protein were generated and named CHO TS cells. It could continue to grow, and the yield of E2 achieved 95.95 mg/L in a tyrosine-free and chemically-defined (CD) medium. Herein, we introduced an alternative to antibiotic-based selections for the establishment of CHO cell lines and provided useful insights for the design and development of CD medium.</span></p></div>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":null,"pages":null},"PeriodicalIF":2.3000,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of bioscience and bioengineering","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1389172323003778","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Efficiently expanding Chinese hamster ovary (CHO) cells, which serve as the primary host cells for recombinant protein production, have gained increasing industrial significance. A significant hurdle in stable cell line development is the low efficiency of the target gene integrated into the host genome, implying the necessity for an effective screening and selection procedure to separate these stable cells. In this study, the genes of phenylalanine hydroxylase (PAH) and pterin 4 alpha carbinolamine dehydratase 1 (PCBD1), which are key enzymes in the tyrosine synthesis pathway, were utilized as selection markers and transduced into host cells together with the target genes. This research investigated the enrichment effect of this system and advanced further in understanding its benefits for cell line development and rCHO cell culture. A novel tyrosine-based selection system that only used PCBD1 as a selection marker was designed to promote the enrichment effect. Post 9 days of starvation, positive transductants in the cell pool approached 100%. Applied the novel tyrosine-based selection system, rCHO cells expressing E2 protein were generated and named CHO TS cells. It could continue to grow, and the yield of E2 achieved 95.95 mg/L in a tyrosine-free and chemically-defined (CD) medium. Herein, we introduced an alternative to antibiotic-based selections for the establishment of CHO cell lines and provided useful insights for the design and development of CD medium.
期刊介绍:
The Journal of Bioscience and Bioengineering is a research journal publishing original full-length research papers, reviews, and Letters to the Editor. The Journal is devoted to the advancement and dissemination of knowledge concerning fermentation technology, biochemical engineering, food technology and microbiology.