期刊
JOURNAL OF BIOTECHNOLOGY
卷 325, 期 -, 页码 360-371出版社
ELSEVIER
DOI: 10.1016/j.jbiotec.2020.09.025
关键词
Cell enrichment; Chinese hamster ovary cells; Clonal variation; Monoclonal antibodies; Recombinant protein production; Silencing RNA
资金
- National Institutes of Health, Phase II STTR grant [R42GM114935]
The novel technology, PTSelect(TM), utilizes siRNA cloned upstream of the gene of interest to enable cell enrichment and selection. Compared to conventional methods, PTSelect(TM) demonstrates comparable stability and productivity, and allows for easy monitoring of productivity changes due to clonal drift.
Currently, stable Chinese hamster ovary cell lines producing therapeutic, recombinant proteins are established either by antibiotic and/or metabolic selection. Here, we report a novel technology, PTSelect (TM) that utilizes an siRNA cloned upstream of the gene of interest (GOI) that is processed to produce functional PTSelect (TM)-siRNAs, which enable cell enrichment. Cells with stably integrated GOI are selected and separated from cells without GOI by transfecting CD4/siRNA mRNA regulated by PTSelect (TM)-siRNAs and exploiting the variable expression of CD4 on the cell surface. This study describes the PTSelect (TM) principle and compares the productivity, doubling time and stability of clones developed by PTSelect (TM) with conventionally developed clones. PTSelect (TM) rapidly established a pool population with comparable stability and productivity to pools generated by traditional methods and can further be used to easily monitor productivity changes due to clonal drift, identifying individual cells with reduced productivity.
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