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. 2008 Mar 21;4(3):e1000039. doi: 10.1371/journal.pgen.1000039

Figure 1. Establishing the lacO/lacI Tethering System.

Figure 1

(A) Map of pCAGASIZ74 encoding myc-lacI -lap2β under the control of the CAG (CMV-chicken actin) promoter. (B) Western blot of fractionated extracts from cells transfected with pCAGASIZ74 and detected with antibody that recognises all isoforms of lap2, - untransfected cells; T- total cell extract; N –nuclear extract; C – cytoplasmic extract; Np – insoluble nuclear extract; Ns – soluble nuclear extract. (C) Western blot with antibody detecting the myc tag on cell lines with lacO sites in chromosome 11 (J1-C3) and stably expressing the myc-tagged lacI-lap2β; J21 = lacO integrated cells without lacI-lap2β, +ve control = transiently transfected cells. (D) Western blot with antibody detecting lacI in the lacO integrated parental B49.5 cells, and in B49.5 and J21 cells stably expressing lacI-lap2β. (E) Immunofluorescence on parental B49.5 lacO integrated cells and B49.5 cells stably expressing lacI-lap2β with antibodies that recognise lacI, myc or lap2. Nuclei are counterstained with DAPI.