MOLECULAR CHARACTERIZATION OF A STABLY TRANSFORMED BOMBYX-MORI CELL-LINE - IDENTIFICATION OF ALTERNATIVE TRANSCRIPTIONAL INITIATION SITES OF THE A3 CYTOPLASMIC ACTIN GENE

We have isolated a stably transformed Bombyx mori cell line containing a novel selectable marker gene, puromycin N-acetyl transferase, under control of transcriptional regulatory signals from the A3 cytoplasmic actin gene. By using this cell line we have identified alternative tr anscriptional initiation sites for the A3 actin gene. One of these sta rt sites is located similar to 35 bp upstream from the previously dete rmined transcription initiation site. The two mRNA start sites are uti lized with a similar efficiencies in the BmN cell line. In addition, w e detected transcripts that initiated in the first intron of the A3 ac tin gene. These transcripts may be synthesised under control of an alt ernative promoter. The stably transformed B. mori cell line used in th is study was also extensively characterized. Integration of the plasmi d molecules into the host genome was demonstrated by Southern and in s itu hybridization analyses. Establishment and characterization of stab ly transformed insect cell lines, like the one described here, represe nts an important step in the development of nonlytic insect expression systems.