Agrobacterium-mediated transformation of safflower and the efficient recovery of transgenic plants via grafting

<p>Abstract</p> <p>Background</p> <p>Safflower <it>(Carthamus tinctorius L.) </it>is a difficult crop to genetically transform being susceptible to hyperhydration and poor <it>in vitro </it>root formation. In addition to traditional uses safflower has recently emerged as a broadacre platform for the production of transgenic products including modified oils and pharmaceutically active proteins. Despite commercial activities based on the genetic modification of safflower, there is no method available in the public domain describing the transformation of safflower that generates transformed T₁progeny.</p> <p>Results</p> <p>An efficient and reproducible protocol has been developed with a transformation efficiency of 4.8% and 3.1% for S-317 (high oleic acid content) and WT (high linoleic acid content) genotypes respectively. An improved safflower transformation T-DNA vector was developed, including a secreted <it>GFP </it>to allow non-destructive assessment of transgenic shoots. Hyperhydration and necrosis of <it>Agrobacterium</it>-infected cotyledons was effectively controlled by using iota-carrageenan, L-cysteine and ascorbic acid. To overcome poor <it>in vitro </it>root formation for the first time a grafting method was developed for safflower in which ~50% of transgenic shoots develop into mature plants bearing viable transgenic T₁seed. The integration and expression of secreted <it>GFP </it>and hygromycin genes were confirmed by PCR, Southern and Western blot analysis. Southern blot analysis in nine independent lines indicated that 1-7 transgenes were inserted per line and T₁progeny displayed Mendelian inheritance.</p> <p>Conclusions</p> <p>This protocol demonstrates significant improvements in both the efficiency and ease of use over existing safflower transformation protocols. This is the first complete method of genetic transformation of safflower that generates stably-transformed plants and progeny, allowing this crop to benefit from modern molecular applications.</p

Development and application of a prototype field test for detection of pre-harvest sprouting in wheat

Established and supported under the Australian Government’s Cooperative Research Centre Progra