Figure 1.

Characterization of human airway epithelial cell lines overexpressing ClC-2. (A) Autoradiograms of genomic DNA from Parental IB3-1 cells and cell lines S1, S2, and S3 digested with NheI (Left) and NheI and NotI (Right) then hybridized with a 0.5-kb SphI-HindIII fragment from the 3′ end of the ClC-2 cDNA. Clones S2 and S3 have intensely hybridizing NheI fragments of 7.5 kb that were digested by NotI to fragments of 4.8 kb and 2.7 kb, consistent with the presence of multiple copies of full-length concatenated plasmids (Lower). Open arrows indicate fragments from the endogenous ClC-2 genes present in each lane. Additional hybridizing signals in some lanes probably represent junction sequences, partial integrations, and/or incomplete digestion. In the diagram, solid boxes and arrows represent the RSV promoter and direction of transcription and hatched boxes indicate the location of the SphI-HindIII probe relative to ClC-2. (B) Autoradiogram of total RNA from Parental IB3-1 cells and clones S1, S2, and S3 (30 μg each) and T84 cells (10 μg) hybridized with a 1.45-kb probe from the 3′ end of the ClC-2 cDNA. Transcripts of 3.3 kb from the endogenous ClC-2 gene are seen in all lanes. S1 has a faint signal at 3.3 kb, comparable to the level of expression in parental cells. S2 and S3 have intense signals at 3.3 and 3.8 kb, indicating expression of stably integrated ClC-2 cDNA. Ethidium bromide staining of the gel indicated that approximately equal amounts of RNA were loaded in each lane (Lower).