SOMATIC AND MEIOTIC CHROMOSOMAL RECOMBINATION BETWEEN INVERTED DUPLICATIONS IN TRANSGENIC TOBACCO PLANTS

Homologous recombination has been extensively studied in bacteria, yea st, and more recently in animal cells, but little is known about this process in plants. We present here an analysis of meiotic and somatic chromosomal recombination between closely linked inverted duplications located on a single chromosomal region in tobacco. Transgenic tobacco lines were constructed by Agrobacterium transformation with plasmid v ectors containing a functional hygromycin phosphotransferase (hyg) sel ectable marker flanked by a pair of defective neomycin phosphotransfer ase (neo) genes positioned as inverted repeats. As each neo gene is mu tated in a different site, recombination between the two defective gen es can be detected following selection for kanamycin-resistant plant c ells. The recombination substrates were designed to allow investigatio n into the nature of molecular events underlying homologous recombinat ion by restriction endonuclease analysis. Chromosomal recombination wa s studied in mitotically dividing cells (cultured leaf mesophyll cells ) and after meiosis (germinated seedlings). Spontaneous somatic recomb inants were recovered at frequencies between approximately 3 x 10(-5) to 10(-6) events per cell. Low dose gamma-irradiation of somatic cells resulted in a threefold maximum increase in the recovery of recombina nts. Recombinants were also detected at low frequency when transgenic T3 seeds were germinated under kanamycin selection. DNA gel blot analy ses demonstrated that homologous recombination occurred mainly as gene conversion unassociated with reciprocal exchange, although a variety of other events including gene coconversion were also observed.