J. Plant Physiol. 160. 931 – 943 (2003)  Urban & Fischer Verlag http://www.urbanfischer.de/journals/jpp Integration, expression and inheritance of transgenes in hexaploid oat (Avena sativa L.) Sophie J. Perret, John Valentine, J. Mike Leggett, Phillip Morris* Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, Wales SY23 3EB, UK Received July 8, 2002 · Accepted November 7, 2002 Summary Two oat varieties, Melys (spring variety) and Bulwark (winter variety) were transformed by particle bombardment of primary embryogenic callus using either a ubi-bar-ubi-gus co-integration vector or co-transformed (Melys) with a ubi-bar plasmid together with one of three plasmids containing the β-glucuronidase (gus) gene under the control of either a rice actin promoter, a CaMV35S promoter or a wheat high molecular weight glutenin promoter. Morphologically normal and fertile transgenic plants were regenerated following callus selection with glufosinate ammonium. Evidence for the integration and functioning of the selectable (bar) and reporter (gus) genes in T o and T 1 plants was confirmed by PCR, Southern hybridisation, fluorescence in situ hybridisation (FISH), histochemical assays, and by progeny analysis. Transformation rates varied from 0.2 to 5.0 lines/plate of callus bombarded, with co-transformation frequencies of 83 to 100%, and co-expression frequencies of 60 to 100 %. Copy numbers for the bar and gus gene varied from 3 to 17and from 2 to 20 respectively. Cell and tissue specific expression of the gus gene was evident from the different promoters, with the HMW glutenin promoter showing endosperm specific expression in T 1 seed. No expression of the gus gene under the CaMV35S promoter was detected in any tissues. Progeny analysis provided evi- dence of Mendelian inheritance of the introduced genes suggesting either one or two unlinked inte- gration sites. This was confirmed by fluorescence in situ hybridisation to chromosome spread prepa- rations. No segregation of the gus gene from the bar gene was observed in any of the progeny derived from co-transformation. Key words: co-transformation – transgene inheritance – transgene expression – oats Abbreviations: GUS = β-glucuronidase. – HMW = high molecular weight. – PAT = phosphinothricin acetyl transferase. – PPT = phosphinothricin Introduction Genetic transformation of many of the important cereals is now well established, and is beginning to provide novel * E-mail corresponding author: phillip.morris@bbsrc.ac.uk breeding material in several species. Rice was the first cereal in which stable transformation and fertile transgenic plants was achieved (Toriyama et al. 1988, Zhang et al. 1988), fol- lowed by maize (Fromm et al. 1990, Rhodes et al. 1988), wheat (Vasil et al. 1992), oats (Somers et al. 1992), and barley (Wan and Lemaux 1994). Successful transformation of oats 0176-1617/03/160/08-931 $ 15.00/0