FORMATION OF AN AUTONOMOUSLY REPLICATING SHUTTLE PLASMID BY IN-VIVO NONHOMOLOGOUS RECOMBINATION

A recombinant plasmid prR1 consisting of a pBR322 fragment and a fragm ent of silkworm DNA was rescued from the F-3 generation of transgenic silkworm Bombyx mori. Plasmid prR1 resulted from nonhomologous recombi nation of the input plasmid pPrC-LTR1.5 with the cell DNA in the F-0 g eneration and additional rearrangements in inheritance. The transgene structure in F-3 is similar to that in F-2 of the same line, where it was also found in an extrachromosomal form and whereform it had been p reviously rescued as plasmid pr8a. Upon pr8a introduction into silkwor m eggs, the newly obtained transgenic F-0 individuals harbored a plasm id identical in restriction mapping to pr8a and prR1 and replicating e xtrachromosomally like the initial one. The prR1 plasmid also retained the ability to replicate in Escherichia coli cells. Sequence analysis of the cell DNA contained in prR1 reveals the presence of two homolog ous elements belonging to a family of evolutionarily conserved frequen t repeats in the silkworm genome.