Construction of TALENs and ZO-1 gene knockout in MDCK I and II cells.

<p>(A) TALEN binding sites in the ZO-1 gene. The left and right arms of TALEN targeting sites are indicated in blue and the spacer region is indicated in red. The initiation codon within the spacer region is highlighted. (B) Immunofluorescence microscopic analysis of ZO-1, ZO-2 and ZO-3 in MDCK II cells transfected with TALEN constructs for ZO-1 gene knockout. After transfection, cells were subcultured on filter inserts for 4 d before analysis. At the boundary of control and ZO-1 knockout cells, characteristic convex curves of cell–cell junctions are observed (<i>arrows</i>). (C) Immunofluorescence microscopic analysis of ZO-1, ZO-2 and ZO-3 in MDCK I cells transfected with TALEN constructs for ZO-1 gene knockout. Similar morphological changes of cell–cell junctions at the boundary of control and ZO-1 knockout cells were observed in MDCK I cells. Staining of ZO-3 was reduced in ZO-1 knockout cells (<i>arrowheads</i>). Scale bars, 10 µm.