T-DNA insertional mutagenesis for functional genomics in rice

We have produced 22 090 primary transgenic rice plants that carry a T-DNA i nsertion, which has resulted in 18 358 fertile lines. Genomic DNA gel-blot and PCR analyses have shown that approximately 65% of the population contai ns more than one copy of the inserted T-DNA. Hygromycin resistance tests re vealed that transgenic plants contain an average of 1.4 loci of T-DNA inser ts. Therefore, it can be estimated that approximately 25 700 taggings have been generated. The binary vector used in the insertion contained the promo terless beta-glucuronidase (GUS) reporter gene with an intron and multiple splicing donors and acceptors immediately next to the right border. Therefo re, this gene trap vector is able to detect a gene fusion between GUS and a n endogenous gene, which is tagged by T-DNA. Histochemical GUS assays were carried out in the leaves and roots from 5353 lines, mature flowers from 70 26 lines, and developing seeds from 1948 lines. The data revealed that 1.6- 2.1% of tested organs were GUS-positive in the tested organs, and that thei r GUS expression patterns were organ- or tissue-specific or ubiquitous in a ll parts of the plant. The large population of T-DNA-tagged lines will be u seful for identifying insertional mutants in various genes and for discover ing new genes in rice.