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Novel drought tolerance genes were identified by screening thousands of random genomic fragments from grass species in transgenic rice. Identification of agronomically important genes is a critical step for crop breeding through biotechnology. Multiple approaches have been employed to identify new gene targets, including comprehensive screening platforms for gene discovery such as the over-expression of libraries of cDNA clones. In this study, random genomic fragments from plants were introduced into rice and screened for drought tolerance in a high-throughput manner with the aim of finding novel genetic elements not exclusively limited to coding sequences. To illustrate the power of this approach, genomic libraries were constructed from four grass species, and screening a total of 50,825 transgenic rice lines for drought tolerance resulted in the identification of 12 reproducibly efficacious fragments. Of the twelve, two were from the mitochondrial genome of signal grass and ten were from the nuclear genome of buffalo grass. Subsequent sequencing and analyses revealed that the ten fragments from buffalo grass carried a similar genetic element with no significant homology to any previously characterized gene. The deduced protein sequence was rich in acidic amino acid residues in the C-terminal half, and two of the glutamic acid residues in the C-terminal half were shown to play an important role in drought tolerance. The results demonstrate that an open-ended screening approach using random genomic fragments could discover trait genes distinct from gene discovery based on known pathways or biased toward coding sequence over-expression.

The Rf-1 gene restores the pollenfertility disturbed by the BT-type malesterile cytoplasm. Nine restrictionfragment length polymorphism (RFLP) markerson chromosome 10, where the Rf-1 geneis located, were converted to PCR-basedmarkers. Genetic analysis was conductedusing the nine markers and a segregatingpopulation consisting of 1042 plants. As aresult, the Rf-1 gene was mappedbetween S12564 Tsp509I and C1361 MwoI. The genetic distance of the twomarker loci was estimated as 0.3 cM. Basedon the map information of the Rf-1gene, it will be possible to introduce alimited chromosomal segment of the Rf-1 region derived from a donor intojaponica rice varieties efficiently andeffectively by marker-assisted selection(MAS). Also, examination of seed purity ofhybrid rice varieties and their parentallines will be facilitated. Furthermore,judging from the genetic distance from theRf-1 locus to S12564 Tsp509Iand C1361 MwoI loci, it seems thatthe Rf-1 gene could be isolated bychromosomal walking from either or both ofthe two marker loci.