Indian Journal of Biotechnology Vol 6, October 2007, pp 504-507 Biolistic-mediated DNA delivery and transient expression of GUS in hypocotyls of Feronia limonia L.—A fruit tree S D Purohit*, S Raghuvanshi 1 and A K Tyagi 1 Plant Biotechnology Laboratory, Department of Botany, Mohanlal Sukhadia University, Udaipur 313 001, India 1 Department of Plant Molecular Biology, University of Delhi South Campus, New Delhi 110 021, India Received 2 May 2006; revised 22 December 2006; accepted 6 March 2007 Feronia limonia L. (Rutaceae) is an important fruit tree of arid horticulture. To be able to introduce genes of interest for its improvement, biolistic-mediated DNA delivery method has been standardized. The hypocotyls explants were bombarded with plasmid pBI121 having gus reporter gene driven by CaMV 35S promoter and npt II under control of nos promoter as selectable marker. The best transient expression of GUS was recorded when tungsten particles coated with DNA were bombarded at a disc pressure of 1350 and 1100 psi from a distance of 9 cm. Some of the bombarded explants are being maintained to obtain viable regenerates for their analyses. Keywords: biolistic, Feronia limonia, transient expression IPC Code: Int. Cl. 8 A01H15/00; C12N15/09, 15/11 Introduction Genetic transformation has become an indispensable tool in plant molecular biology and functional genomics research 1 . Since its initial application to plants 2 , more than 144 plant species have been successfully transformed—representing almost all major phylogenetic lineages of the plant kingdom 3 . In trees, where long regeneration cycles, high background polymorphism, large sizes and outcrossing systems of mating make the production of near-isogenic lines and large experiments difficult, manipulation at the cellular and molecular level is an attractive alternative for establishing direct linkages between genes and adaptively significant phenotypes 1 . Successful development of transgenics in woody trees has been primarily limited by lack of efficient in vitro regeneration protocols and suitable gene transfer mechanism. Currently, it is possible to regenerate successfully many of the woody species which were initially considered to be recalcitrant 4-9 . Agrobacterium and biolistic are the most commonly employed gene delivery systems but physical nature of particle bombardment can potentially overcome many of the biological barriers, such as host-range specificity associated with Agrobacterium 10 . Transformation of tree species via particle bombardment was achieved in Carica papaya 11 and a hybrid poplar 12 . Reports on biolistic-mediated gene transfer in tree species have been reviewed by Khurana and Khurana 13 , Busov et al 1 and Petri and Burgos 14 . Feronia limonia L. (Family: Rutaceae) is an important fruit tree suitable for cultivation under semi- arid conditions and generally recommended for use in reforestation and wasteland reclamation projects 15 . In vitro regeneration protocol for F. limonia through callus organogenesis 16 , adventitious bud formation on hypocotyls segments 17 and cotyledonary and mature node segments 16 have been standardized in authors’ laboratory. In the present communication, authors report, for the first time, the transient expression of β- glucuronidase (gus) reporter gene through biolistic delivery in hypocotyls segments of F. limonia. Materials and Methods Seeds of F. limonia obtained from fresh ripe fruits were surface sterilized with 0.1% mercuric chloride (HgCl 2 ) for 5 min and rinsed 3-4 times them with sterilized double-distilled water. Seeds were then inoculated on water-agar (0.8%). Germination started within 7-8 d. Fully-grown seedlings having an average hypocotyls length of 3.0 cm were used to obtain explants. Regeneration medium 17 contained MS salts plus 2.0 mg L -1 BAP and 0.5 mg L -1 Kn. Sorbitol and mannitol @ 3.0% were incorporated in ________________ *Author for correspondence: Tel & Fax: 91-294-2410300 E-mail: sdp_56@yahoo.com/hotmail.com