Abstract:

In this study, we constructed a super binary vector to evaluate the potential of the twin T-DNA system for generating selectable marker-free progeny plants in chrysanthemum plants. The first T-DNA of the vector, contains the hygromycin phosphotransferase (hpt) selectable gene, while the second T-DNA, bears the β-glucuronidase gene (gus), featuring the gene of interest. The two T-DNA regions were located adjacent to each other with no intervening region. 506 resistant chrysanthemum plants were then obtained by Agrobacterium- mediated transformation with this vector, analysis of transgene inheritance was facilitated by PCR amplification and Southern blot from leaf tissue, the primary co-transformation frequency was 38.4%. A total of seventeen hpt-resistant/gus-active T0 plants were evaluated for segregation in the next generation, and among these, approximately 15.8% had transgene inserts which segregated in the T₁ progeny to yield chrysanthemum plants without selectable marker gene. Overall, the twin T-DNA systems appeared to be a useful approach to generate marker free transgenic plants, thereby eliminating public concerns regarding proliferation of antibiotic and herbicide resistance genes in the environment.

Key words: chrysanthemum, transgenic, twin T-DNA, co-transformation, marker-free