T‐DNA insertional mutagenesis for functional genomics in rice

Top Cited Papers
Open Access
Publisher Website
Google Scholar
Abstract
Summary: We have produced 22 090 primary transgenic rice plants that carry a T‐DNA insertion, which has resulted in 18 358 fertile lines. Genomic DNA gel‐blot and PCR analyses have shown that approximately 65% of the population contains more than one copy of the inserted T‐DNA. Hygromycin resistance tests revealed that transgenic plants contain an average of 1.4 loci of T‐DNA inserts. Therefore, it can be estimated that approximately 25 700 taggings have been generated. The binary vector used in the insertion contained the promoterless β‐glucuronidase (GUS) reporter gene with an intron and multiple splicing donors and acceptors immediately next to the right border. Therefore, this gene trap vector is able to detect a gene fusion between GUS and an 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 7026 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 their GUS expression patterns were organ‐ or tissue‐specific or ubiquitous in all parts of the plant. The large population of T‐DNA‐tagged lines will be useful for identifying insertional mutants in various genes and for discovering new genes in rice.