Somatic embryogenesis and Agrobacterium-mediated transformation of Gladiolus hybridus cv. ‘Advance Red’

Autor: Jian Wu, Junna He, Chao Liu, Benhe Gong, Chen Liu, Shanshan Seng, Xionghui Zhong, Chenyu Wu, Mingfang Yi, Juanjuan Sui, Muhammad Ali Khan
Rok vydání: 2014
Předmět:
Zdroj: Plant Cell, Tissue and Organ Culture (PCTOC). 120:717-728
ISSN: 1573-5044
0167-6857
DOI: 10.1007/s11240-014-0639-5
Popis: Stable and efficient Agrobacterium-mediated transformation of Gladiolus hybridus cv. ‘Advance Red’ was established via a somatic embryogenesis regeneration system in this study. Treatment with N-phenyl-N′-1, 2, 3-thiadiazol-5-ylurea was sufficient to induce embryogenic callus and somatic embryogenesis in G. hybridus ‘Advance Red’. Cormel slices were used as explants and pre-cultured for 5 days prior to selection culture with 9.48 μM hygromycin and 0.52 mM cefotaxime. Treated slices were inoculated with Agrobacterium stain GV3101 harboring a binary vector, pCAMBIA1301, containing the β-glucuronidase (gus) reporter and hygromycin phosphotransferase genes (hpt). 10 min of infection was determined as the optimal time for transformation. Compared with the normal 3-day co-cultivation period, prolonged co-culture for 12 days led to significantly higher transient gus expression rate (50.52 ± 2.83 %). Permanent expression of gus was observed in leaves, roots and whole plantlets. T-DNA insertion was further confirmed using PCR and Southern blot analyses. Our data collectively indicate that extended pre-cultivation and co-cultivation improve transformation efficiency, and the somatic embryogenic regeneration system effectively facilitates the transformation of G. hybridus. This newly developed transformation system may be successfully applied for functional analysis of genes and quality modifications in G. hybridus.
Databáze: OpenAIRE
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