GENETICS AND GENOMICS OF GRAPE GROWTH, DEVELOPMENT, AND QUALITY
Location: Grape Genetics Research
Title: Influences of Agrobacterium rhizogenes strains, plant genotypes, and tissue types on the induction of transgenic hairy roots in Vitis species
Submitted to: Vitis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 31, 2011
Publication Date: August 15, 2011
Citation: Jittayasothorn, Y., Chen, S., Wang, X., Zhong, G. 2011. Influences of Agrobacterium rhizogenes strains, plant genotypes, and tissue types on the induction of transgenic hairy roots in Vitis species. Vitis. 50(3):107-114.
Agrobactrium rhizogenes can infect various types of plant tissues and induce development of “hairy roots” from these tissues in a few weeks. These “hairy roots” provide an excellent tool for high throughput functional genomics research in which functionalities of genes can be evaluated quickly. A. rhizogenes-induced hairy roots have been demonstrated in grapevines, but the induction was carried out in only a few species and genotypes. There are many factors, such as agrobacterium strains, plant genotypes and tissue types for inoculation, which can impact the efficiencies of hairy root induction. In this study, we evaluated the influences of 3 A. rhizogenes strains (15384, A4, and K599), 4 different tissue types (internode, stem, and petiole on and young shoots excised from in vitro plants) and 14 grape species on the induction of hairy roots. Our results revealed that both 15834 and A4 strains, but not K599, were effective in inducing hairy roots in grape species and different grape species and tissue types responded differently to the induction. Among the 14 species evaluated, V. cinerea, V. labrusca, V. treleasii, and V. vinifera appeared to be much more responsive to hairy root induction than others when induction was carried out on in vitro plants. Internode tissue, compared with stem and petiole tissues, had a higher frequency of hairy root induction. However, young shoots excised from in vitro plants were found to be the most suitable tissue for hairy root induction, better than internode, stem or petiole tissue on in vitro plants. Among the six species, V. cinerea, V. ficifolia, V. girniana, V. jacquemontii, V. palmata, and V. vinifera, evaluated with excised young shoots, V. cinerea and V. palmata were most responsive to hairy root induction. The results obtained from this study furthered our understanding of the influences of various factors on hairy root induction in grapevines. We identified several species which are highly responsive to hairy induction and can be useful for grapevine functional genomics research in the future.
Agrobacterium rhizogenes-mediated induction of transgenic hairy roots was previously demonstrated in Vitis vinifera L. and a few other Vitis species. In this study, 13 Vitis species, including V. aestivalis, V. afghanistan, V. champinii, V. doaniana, V. flexuosa, V. labrusca, V. nesbittiana, V. palmata, V. piasezkii, V. thunbergii, V. treleasii, V. vinifera L, and V. xnovae angelae were screened for hairy root induction by co-cultivating in vitro plants with virulent Agrobacterium rhizogenes 15834, A4, or K599 harboring binary vector pBIN61-EGFPHA, which produces green fluorescent protein signal for visual examination of transgenic hairy roots. Frequencies of callus formation and transgenic hairy root induction were recorded daily for 2 weeks after in vitro plants had been inoculated. The results showed that V. labrusca (var. ‘Concord’) had the highest frequency of transgenic callus and hairy root induction. Hairy roots were also successfully induced for V. treleasii, V. champinii, V. cinerea, and V. vinifera, but their frequencies were much lower than that for V. labrusca. In contrast, no hairy root was generated from other species during the 2-week observation period. In addition, we found that the developmental stage or age of plant material also played an important role in transgenic hairy root induction. Our results showed that hairy roots could be induced by 15834 and A4 strains with similar transformation efficiencies, however, K599 was found not to be suitable for hairy root induction in the Vitis species. In general, the transgenic hairy roots appeared to be bright white and much thicker than nontransgenic roots and they could grow up to 2-cm long in one-week period on hairy root culture medium. These results provided important addition to the knowledge of transgenic hairy root induction in the Vitis species.