Highly efficient plant regeneration and Agrobacterium-mediated transformation of Helianthus tuberosus L

• Published in: Industrial crops and products Vol. 83; pp. 670 - 679
• Main Authors: Kim, Mee-Jin, An, Dong-Ju, Moon, Ki-Beom, Cho, Hye-Sun, Min, Sung-Ran, Sohn, Jung-Hoon, Jeon, Jae-Heung, Kim, Hyun-Soon
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 01.05.2016
• Subjects: Agrobacterium tumefaciens; Helianthus tuberosus; Jerusalem artichoke; Regenerated plants; Shoot regeneration; Stable transformation; KIN; Stable transformation; Shoot regeneration; IAA; Regenerated plants; Jerusalem artichoke; PCR; BA; NAA
• ISSN: 0926-6690; 1872-633X
• DOI: 10.1016/j.indcrop.2015.12.054

•Shoot regeneration efficiency from regenerated shoots was over 50%.•MS medium supplemented with 1.0mgL−1 zeatin was the best for shoot regeneration.•The regeneration frequency was higher under dark conditions than under the 16L/8D.•An efficient Agrobacterium-mediated transformation system was successfully achieved.•This method will be useful for genetic improvement of H. tuberosus. In our current study, to develop an efficient regeneration and transformation system of Helianthus tuberosus, we verified effects of plant growth hormones, growth conditions, explant type, and transformation conditions. Leaf segments from regenerated shoots showed higher regeneration efficiency on MS basal medium containing 1.0mgL−1 zeatin under darkness than those from maintained in vitro plant. To carry out transformation, various parameters including plant materials, Agrobacterium cell density, immersion time, and Agrobacterium strains (leaf segment, OD600-0.6, 60min, and Agrobacterium tumefaciens LBA4404 harboring binary vector pCAMBIA1301) have been determined. The putatively transformed H. tuberosus were screened by survival rate and beta-glucuronidase (GUS) histochemical assay following two cycles of 3.0mgL−1 hygromycin selection at callus stage. The presence of the selectable marker gene hygromycin phosphotransferase and the GUS reporter gene with intron was then confirmed by genomic PCR, Southern blot, reverse transcriptase PCR (RT-PCR) and GUS histochemical assay. The method presented here could be helpful in genetic improvement of H. tuberosus through efficient shoot regeneration and stable Agrobacterium-mediated transformation.