Large-scale production of enhancer trapping lines for rice functional genomics

Using a rapid and efficient Agrobacterium tumefaciens mediated rice transformation protocol, we had got about 100,000 independent enhancer trapping lines of japonica rice variety Nipponbare. T, seeds had already been obtained from 60,000 lines. Regeneration of transgenic plants could be as short as 3 months starting from callus induction, thus the Occurrence of somaclonal variation was reduced to a minimum level. The frequencies of hygromycin-resistant callus induction and plant regeneration were over 90 and 80%, respectively. PCR and Southern blot analysis showed that about 95% hygromycin resistant plants were transgenic. Southern blot and segregation analysis of T, seeds showed that about 40% transgenic plants contained single copy insertion, and the average copy number was 1.7. Histochemical assay of To plants revealed expression patterns in root, leaf, embryo and endosperm of the reporter gene beta-glucurodinase (GUS). This large-scale enhancer trapping pool provides a valuable tool for functional analysis of rice genome. (C) 2004 Elsevier Ireland Ltd. All rights reserved.