Inheritance of an induced male-sterile trait in transgenic plants expressing an engineered unedited atp9 mitochondrial gene

 Flowers of tobacco transformed with the genomic (unedited) coding sequence of the mitochondrial atp9 gene (u-atp9), fused to the yeast coxIV mitochondrial targeting pre-sequence, exhibited either semi-fertile or male-sterile phenotypes. The inheritance of the induced male-sterile trait, was first investigated on a population of 25 plants of the R0 generation and then on R1, F1 and R2 progeny of the lines H2.11 and H2.16. The analyses showed that the male-sterile character was generally inherited as a Mendelian trait for one or two loci, and that u-atp9 is involved in the male-sterile phenotype and is stably inherited in the progeny of the original transformants. Molecular investigations performed on the plants of the R0, R1 and R2 generations confirmed the genetic analyses. The final proof of the involvement of u-atp9 in the emergence of the male-sterile trait was provided from experiments dealing with the restoration of male fertility. By using antisense RNA technology, we showed that crossing male-sterile plants containing the u-atp9 transgene with transgenic plants containing the same gene in antisense orientation (as-atp9) produces F1 progeny restored to male fertility. The high expression of as-atp9 causes a dramatic reduction of u-atp9 transcript levels and suppresses their deleterious effects, resulting in normal flower development and seed production.