Agrobacterium-mediated transformation of sunflower (Helianthus annus L.):: A simple protocol

A simple and reproducible Agrobacterium tumefaciens-mediated transformation system was developed for sunflower (Helianthus annuus L.) 'KBSH-1'. The objective was to substantially eliminate the in vitro regeneration component from the transformation protocol. Two-d-old seedlings with one cotyledon detached were infected with Agrobacterium tumefaciens strain LBA 4404/pKIWI105 harbouring genes for beta-glucuronidase (GUS) and neomycin phosphotransferase (NPT II). Following co-cultivation, seedlings were grown aseptically for 5d on a growth regulator-free basal medium supplemented with 250 mu g ml(-1) cefotaxime (a bacteriostat used to control gram negative bacteria). Seedlings were screened for transient GUS expression and the shoot portions of the putative transformants were utilized for propagation as transgenic plants. The excised shoots that initiated roots following selection were subsequently transferred to a glasshouse. Molecular analysis of transgenic plants confirmed concordance between the presence of foreign genes and enzyme activity. The transformation regime facilitated rapid generation of up to 2% phenotypically normal fertile plants containing functional transgenes. The transmission and integration of the marker genes in the progeny is demonstrated. (C) 1999 Annals of Botany Company.