Genomic Southern blot analysis

Genomic DNAs (20 ^g each) isolated from each CHO cell line were digested with SspI. A DIG-labeled probe was prepared using the PCR DIG Probes Synthesis Kit (Roche Diagnostics, Mannheim, Germany). The labeling by PCR was carried out with primer pairs (5'-AAGGACATCCCCC ATCAACACAATG-3' and 5'-AGGGGCCAGTCTTAGTCAAACCACC-3') according to the manufacturer's instructions. Each DNA sample was electrophoresed and transferred to Hybond N+ (Amersham Biosciences, Piscataway, NJ). The membrane was hybridized with the Dig-labeled probe in DIG Easy Hyb (Roche Diagnostics) overnight at 42°C. Then the blot was washed with a solution of 2 x SSC and 0.1% SDS for 10 minutes at room temperature, and with a solution of 0.1 x SSC and 0.1% SDS for 15 minutes at 65 °C. The signals were visualized by the DIG system (Roche Diagnostics).

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