Abstract
Chinese hamster ovary (CHO) cells [1] are today a very important host for the commercial-scale production of protein pharmaceuticals. Two sub clones of CHO cells, proline-requiring CHO K1 [2] and the dihydrofolate reductase (DHFR) gene-deficient CHO DG44 [3], are the most widely used for both scientific research and industrial applications [4][5,6]. Previously, we constructed a genomic bacterial artificial chromosome (BAC) library from mouse Dhfr-amplified CHO DR1000L-4N cell genome, which was provided 5-fold coverage of the CHO cell genome and analyzed the structure of amplicons of exogenous Dhfr amplification [7]. The BAC clones of this library could be landmarks for a physical map for CHO cell genome that are essential to the basic research and industrial application of CHO cell genome. In this study, we constructed the detail chromosomal physical map of CHO DG44 cell and investigated the chromosome rearrangements among CHO K1, DG44, and primary Chinese hamster cells. Moreover, to determine the effect of the palindrome structure on Dhfr amplification in CHO cells, we constructed three types of expression vectors with or without the junction region of the proposed amplicon and investigated the gene amplification and expression levels in transfected CHO DG44 cells.
Highlights
Chinese hamster ovary (CHO) cells [1] are today a very important host for the commercial-scale production of protein pharmaceuticals
Construction of bacterial artificial chromosome (BAC)-based physical map for Chinese hamster ovary cells A CHO genomic BAC library consisted of 122,281 clones was constructed in a previous study [7]
Three hundred BAC clones were randomly selected from this library and mapped onto each chromosome of CHO DG44 cells by BAC-FISH
Summary
Chinese hamster ovary (CHO) cells [1] are today a very important host for the commercial-scale production of protein pharmaceuticals. Two sub clones of CHO cells, proline-requiring CHO K1 [2] and the dihydrofolate reductase (DHFR) gene-deficient CHO DG44 [3], are the most widely used for both scientific research and industrial applications [4][5,6]. The BAC clones of this library could be landmarks for a physical map for CHO cell genome that are essential to the basic research and industrial application of CHO cell genome. We constructed the detail chromosomal physical map of CHO DG44 cell and investigated the chromosome rearrangements among CHO K1, DG44, and primary Chinese hamster cells. To determine the effect of the palindrome structure on Dhfr amplification in CHO cells, we constructed three types of expression vectors with or without the junction region of the proposed amplicon and investigated the gene amplification and expression levels in transfected CHO DG44 cells
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