Describe the process of developing a stable CHO cell line for therapeutic protein production.

CHO (Chinese Hamster Ovary) cell line development involves: 1) Vector design - expression cassette with strong promoter (CMV, EF1a), gene of interest, selection marker (DHFR, GS), and regulatory elements. 2) Transfection - introduce DNA via electroporation, lipofection, or nucleofection. 3) Selection pressure - culture in selective media (methotrexate for DHFR, methionine sulfoximine for GS) to enrich stable integrants. 4) Amplification - increase selection pressure to amplify gene copies and expression. 5) Clone screening - single-cell cloning by limiting dilution, FACS, or ClonePix; screen hundreds of clones for productivity, growth, and stability. 6) Clone selection - evaluate top clones for specific productivity (pg/cell/day), product quality attributes, and genetic stability. 7) Cell banking - create Master and Working Cell Banks. Timeline: 6-12 months for high-producing stable pools, longer for clonal cell lines.