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Iwaki T, Castellino FJ.
A single plasmid transfection that offers a significant advantage associated
with puromycin selection in Drosophila Schneider S2 cells expressing
heterologous proteins.
Cytotechnology. 2008 May;57(1):45-9.
W.M. Keck Center for Transgene Research, University of Notre Dame, 235 Raclin-Carmichael
Hall, Notre Dame, IN, 46556, USA, tiwaki@nd.edu.
The Drosophila Schneider S2 (S2) Expression System enables expression of
recombinant proteins constitutively, as well as inductively. This system can
establish both transient and stable transformants with various selection
markers. The generation of stable cell lines for increased expression or large
scale expression of the desired protein is currently accomplished by
cotransfection of both the expression and selection vectors. The selection
vectors, pCoHYGRO and pCoBLAST, are commercially available using hygromycin-B
and blasticidin S, respectively. Recently, we generated a plasmid, pCoPURO, for
selection of transfected S2 cells using puromycin, which allows
significant acceleration of the selection time. Although co-transfection of the
selection marker with the plasmid for heterologous protein expression is
functional in stable expression at short culture periods, the expression levels
of stable transformants are continuously decreased during long culture times. To
overcome this limitation, we generated pMT-PURO, a new plasmid that contains
both the expression cassette and puromycin selection marker in a single
plasmid. This system allows rapid selection and maintenance of the transformed
S2 lines for extended culture periods. PMID: 19003171 |
