Development of stable cell lines for production or regulated expression using matrix attachment regions.
One of the major hurdles of isolating stable, inducible or constitutive high-level producer cell lines is the time-consuming selection procedure. Given the variation in the expression levels of the same construct in individual clones, hundreds of clones must be isolated and tested to identify one or more with the desired characteristics. Various boundary elements (BEs), matrix attachment regions, and locus control regions (LCRs) were screened for their ability to augment the expression of heterologous genes in Chinese hamster ovary (CHO) cells. Of the chromatin elements assayed, the chicken lysozyme matrix-attachment region (MAR) was the only element to significantly increase stable reporter expression. We found that the use of the MAR increases the proportion of high-producing clones, thus reducing the number of clones that need to be screened. These benefits are observed both for constructs with MARs flanking the transgene expression cassette, as well as when constructs are co-transfected with the MAR on a separate plasmid. Moreover, the MAR was co-transfected with a multicomponent regulatable beta-galactosidase expression system in C2C12 cells and several clones exhibiting regulated expression were identified. Hence, MARs are useful in the development of stable cell lines for production or regulated expression.
Zahn-Zabal
M
M
Laboratory of Molecular Biotechnology, Center for Biotechnology UNIL-EPFL, University of Lausanne, CBUE, DC-IGC, CH-1015, Lausanne, Switzerland.
Kobr
M
M
Girod
P A
PA
Imhof
M
M
Chatellard
P
P
de Jesus
M
M
Wurm
F
F
Mermod
N
N
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D002645:Chickens; D002843:Chromatin / Q000235:genetics; D006224:Cricetinae; D005109:Extracellular Matrix / Q000378:metabolism*; D005786:Gene Expression Regulation; D009113:Muramidase / Q000235:genetics* / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D014162:Transfection; D019076:Transgenes
D002843:Chromatin; D009113:Muramidase