Chin Bull Bot ›› 2016, Vol. 51 ›› Issue (3): 363-368.doi: 10.11983/CBB15102

• TECHNIQUES AND METHODS • Previous Articles     Next Articles

Induction of Hairy Roots of Dianthus chinensis and Its Plant Regeneration

Heping Shi*, Bei Wang, Shunan Yang, Yapeng Guo   

  1. Guangdong Provincial Key Laboratory of Biotechnology for Plant Development, School of Life Sciences, South China Normal University, Guangzhou 510631, China
  • Received:2015-06-06 Accepted:2015-09-28 Online:2016-05-24 Published:2016-05-01
  • Contact: Shi Heping
  • About author:

    ? These authors contributed equally to this paper


This study established an efficient system for inducing hairy roots and plant regeneration of Dianthus chinensis. Hairy roots could be induced from the basal surface of leaf explants of D. chinensis at 10 days after inoculation with the strain of Agrobacterium rhizogenes ATCC15834 harbouring the wild agropine-type plasmid. The percentage of rooting leaf explants was 95% at 30 days after inoculation. Hairy roots could grow rapidly and autonomously in liquid or solid plant-growth regulator-free MS medium. The transformation was confirmed by PCR amplification of the rol gene of the Ri plasmid from D. chinensis hairy roots. Hairy roots could form light green callus after culture on MS + 2.0 mg·L-1 6-BA + 0.2 mg·L-1 NAA for 15 days. Adventitious shoots were gradually produced from calli after culture on MS + 1.0 mg·L-1 6-BA + 0.02 mg·L-1 NAA for 30 days. Compared to the control, pot-grown plants regenerated from hairy roots had shorter internodes and flowering was earlier by 18 days.

Table 1

Primer sequences used for the PCR amplification"

Target gene Primer sequence
rol B

rol C

Fig. 2

Induction and in vitro culture of hairy roots and its plant regeneration of Dianthus chinensis ((A) Hairy root formation from leaf explants infected by the strain of Agrobacterium rhizogenes ATCC15834 for 12 days; (B) Solid medium culture of hairy roots for 14 days; (C) Liquid medium culture of hairy roots for 14 days; (D) Calli formation from hairy roots cultured on MS+2.0 mg·L-1 6-BA+0.2 mg·L-1 NAA for 15 days; (E) Adventitious shoots formation from calli after cultured on MS+1.0 mg·L-1 6-BA+0.02 mg·L-1 NAA for 20 days; (F) Multiplication of adventitious shoots on MS+0.1 mg·L-1 6-BA+0.2 mg·L-1 NAA; (G) Pot-grown untransformed (control) plants of Dianthus chinensis for 45 days; (H) Pot-grown regenerated plants from hairy roots for 45 days)"

Fig. 2

Gel electrophoresis analysis of PCR fragments of rol B and rol C genes amplified from the genome DNA of ha- iry roots(Lane 1: 1 kb DNA marker; Lane 2-5: Fragments with rol B primers;Lane 6-9: Fragments with rol C primers; Lane 2 and 6: Fragments amplified from the colony of Agrobacterium rhizogenes ATCC15834; Lane 3 and 7: Fragments from genome DNA of untransformed roots; Lane 4, 5, 8, and 9: Fragments amplified from genome DNA of hairy roots)"

Table 2

Comparison of growth morphology of untransformed and hairy root-regenerated plants of Dianthus chinensis"

Control plants Hairy roots regenerated plants
Plant height (cm) 22.50±2.64 17.20±2.25
Internode Length of 1st leaf from apex (cm)
Internode Length of 2nd leaf from apex (cm)
Internode Length of 3rd leaf from apex (cm)
Length of 1st leaf from apex (cm)
Length of 2nd leaf from apex (cm)
Length of 3rd leaf from apex (cm)
Width of 1st leaf from apex (cm)
Width of 2nd leaf from apex (cm)
Width of 3rd leaf from apex (cm)
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