A new method for the identification and isolation of genes essential for Arabidopsis thaliana seed germination

Seed viability, dormancy and germination efficiency are very important aspects of the life cycle of plants and their potential to survive and spread in the environment. To characterize the genes controlling these processes, we have devised a technique for the selection of mutants impaired in seed germination. Selection for such a trait is complicated by physiological factors that interact with these processes and affect seed germination efficiency. The distinction between low seed germination potential due to physiological factors that interfere with seed maturation or germination and germination deficiency due to genetic factors was based on screening for tagged mutations. Arabidopsis thaliana T-DNA primary transformants obtained by an in planta transformation technique are all heterozygotes. We screened for lack of germination of 1/4 of the seeds in the progeny of independent transformants, and simultaneously for the abnormal segregation (2:1 instead of 3:1) of a kanamycin resistance marker carried by the T-DNA inserted into the genome of these primary transformants in the plants that germinate. This yielded several mutants affected in the germination processes. One of the mutants, designated ABC33, was further characterized. Once the viable embryos from non-germinating seeds were removed from their testa, they grew and displayed a dwarf phenotype which could be complemented by providing gibberellic acid. A genetic and molecular analysis, based on the characterization of the flanking genomic sequences of the T-DNA insert, showed that ABC33 is a new loss-of-function allele at the GA1 locus.