In Vitro Cellular & Developmental Biology - Plant

, Volume 54, Issue 6, pp 606–611 | Cite as

In vitro preservation and micropropagation of Oreocharis mileense (W.T. Wang) M. Möller & A. Weber (Gesneriaceae) through shoot organogenesis

  • Dandan Wang
  • Xinhui Li
  • Zhiying Cheng
  • Chunlin LongEmail author


Oreocharis mileense (W.T. Wang) M. Möller & A. Weber is endemic to China and was considered to be extinct because it had not been seen in the wild since the first collection in 1906. In 2006, the species was rediscovered in Shilin County, Yunnan Province. Oreocharis mileense was considered critically endangered for its narrow geographic range and extremely small population. An efficient method to preserve plant germplasm by in vitro culturing of O. mileense has not been reported. In this study, an orthogonal array with three factors (6-benzyladenine, BA; α-naphthaleneacetic acid, NAA; and sucrose), at four levels was performed, and shoot induction as well as shoot proliferation were recorded. The results were analyzed to determine the most significant components and the optimum combination for micropropagation of O. mileense. The results showed that: (1) organogenesis was easily induced by different combinations of plant-growth regulators and sucrose; (2) NAA and sucrose had the most significant effect on shoot induction and shoot multiplication, and (3) the optimum induction and proliferation media were 0.5 mg L−1 BA + 0.2 mg L−1 NAA + 30 g L−1 sucrose and 1 mg L−1 BA + 0.1 mg L−1 NAA + 30 g L−1 sucrose, respectively.


Oreocharis mileense Rediscovery Critically endangered In vitro preservation Micropropagation 



The authors in particular wish to thank Professor David E. Boufford for editing the English.

Funding information

This study was supported by the Ministry of Science and Technology of China (2005DKA21006, 2012FY110300), the National Natural Science Foundation of China (31761143001, 31870316, 31070288, 31560063), the Ministry of Education of China (yldxxk201819, 2015MDTD16C, B08044), and the Key Laboratory of Ethnomedicine (Minzu University of China) of the Ministry of Education of China (KLEM-ZZ201806).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Supplementary material

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11627_2018_9941_MOESM2_ESM.docx (345 kb)
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  1. Cardoso JC, Habermann G (2014) Adventitious shoot induction from leaf segments in Anthurium andreanum is affected by age of explant, leaf orientation and plant growth regulator. Hortic Environ Biotechnol 55:56–62CrossRefGoogle Scholar
  2. Chen WH, Shui YM, Yang JB, Wang H, Nishii K, Wen F, Zhang ZR, Möller M (2014) Taxonomic status, phylogenetic affinities and genetic diversity of a presumed extinct genus, Paraisometrum W.T. Wang (Gesneriaceae) from the karst regions of Southwest China. PLoS ONE 9:e107967CrossRefGoogle Scholar
  3. Corral P, Mallón R, Rodríguez-Oubiña J, González ML (2011) Multiple shoot induction and plant regeneration of the endangered species Crepis novoana. Plant Cell Tissue Organ Cult 105:211–217CrossRefGoogle Scholar
  4. Djilianov D, Genova G, Parvanova D, Zapryanova N, Konstantinova T, Atanassov A (2005) In vitro culture of the resurrection plant Haberlea rhodopensis. Plant Cell Tissue Organ Cult 80:115–118CrossRefGoogle Scholar
  5. Gao Q, Xu WB (2011) Paraisometrum W. T. Wang, a newly recorded genus of Gesneriaceae from Guizhou, China. Acta Botan Boreali-Occiden Sin 31:858–860 (In Chinese)Google Scholar
  6. Geier T (1986) Factors affecting plant regeneration from leaf segments of Anthurium scherzerianum Schott (Araceae) cultured in vitro. Plant Cell Tissue Organ Cult 6:115–125CrossRefGoogle Scholar
  7. Li J, Xing Q, Chen WL, Guo DH, Shi L (2009) Tissue culture and rapid propagation of Chirita medica D. Fang ex W.T. Wang. Propag Ornam Plant 9:97–101Google Scholar
  8. Li AH, Wang DD, Yu BZ, Yu XM, Li WQ (2014) Maintenance or collapse: responses of extraplastidic membrane lipid composition to desiccation in the resurrection plant Paraisometrum mileense. PLoS One 9:e103430CrossRefGoogle Scholar
  9. Lin L, Yuan B, Wang DD, Li WQ (2014) Cryopreservation of adventitious shoot tips of Paraisometrum mileense by droplet vitrification. CryoLetters 35:22–28PubMedGoogle Scholar
  10. Liu BL, Yang X, Liu J, Dong Y, Wang YZ (2014) Characterization, efficient transformation and regeneration of Chirita pumila (Gesneriaceae), a potential evo-devo model plant. Plant Cell Tissue Organ Cult 118:357–371CrossRefGoogle Scholar
  11. Lloyd G, McCown B (1980) Commercially-feasible micropropagation of mountain laurel, Kalmia latifolia, by use of shoot-tip culture. Int Plant Propagators Soc 30:421–427Google Scholar
  12. Lo KH (1997) Factors affecting shoot organogenesis in leaf disc culture of African violet. Sci Hortic 72:49–57CrossRefGoogle Scholar
  13. Ma GH, He CX, Ren H, Zhang QM, Li SJ, Zhang XH, Eric B (2010) Direct somatic embryogenesis and shoot organogenesis from leaf explants of Primulina tabacum. Biol Plant 54:361–365CrossRefGoogle Scholar
  14. Ma GH, Teixeira da Silva JA, Lü JF, Zhang XH, Zhao JT (2011) Shoot organogenesis and plant regeneration in Metabriggsia ovalifolia. Plant Cell Tissue Organ Cult 105:355–361CrossRefGoogle Scholar
  15. Ma YP, Chen G, Grumbine RE, Dao ZL, Sun WB, Guo HJ (2013) Conserving plant species with extremely small populations (PSESP) in China. Biodivers Conserv 22:803–809CrossRefGoogle Scholar
  16. Mithila J, Hall J, Victor JMR, Saxena P (2003) Thidiazuron induces shoot organogenesis at low concentrations and somatic embryogenesis at high concentrations on leaf and petiole explants of African violet (Saintpaulia ionantha Wendl.). Plant Cell Rep 21:408–414CrossRefGoogle Scholar
  17. Möller M, Forrest A, Wei YG, Weber A (2011) A molecular phylogenetic assessment of the advanced Asiatic and Malesian didymocarpoid Gesneriaceae with focus on non-monophyletic and monotypic genera. Plant Syst Evol 292:223–248CrossRefGoogle Scholar
  18. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  19. Padmanabhan P, Murch SJ, Sullivan JA, Saxena PK (2015) Micropropagation of Primulina dryas (Dunn) Mich. Möller & A. Webber: high frequency regeneration from leaf explants. Sci Hortic 192:250–255CrossRefGoogle Scholar
  20. Shui YM (2006) Botanists rediscover a plant thought to be extinct. Bull Chin Acad Sci 20:206–207 (In Chinese)Google Scholar
  21. Shui YM, Cai J (2007) “100-years-lost” plant re-emerges in Yunnan, China. Samara 12:3Google Scholar
  22. Taha RM, Daud N, Hasbullah NA, Awal A (2009) Somatic embryogenesis and production of artificial seeds in Sintpaulia ionantha Weddl. Acta Hotic 829:331–336Google Scholar
  23. Tan Y, Wang Z, Sui XY, Hu GW, Motley T, Long CL (2011) The systematic placement of the monotypic genus Paraisometrum (Gesneriaceae) based on molecular and cytological data. Plant Divers 33:465–476Google Scholar
  24. Wang WT, Pan KY, Li ZY (1997) New taxa, new combinations, and notes on Chinese Gesneriaceae. Novon 7:423–435CrossRefGoogle Scholar
  25. Xiao LH, Yang G, Zhang LC, Yang XH, Zhao S, Ji ZZ, Zhou Q, Hu M, Wang Y, Chen M, Xu Y, Jin HJ, Xiao X, Hu GP, Bao F, Hu Y, Wan P, Li LG, Deng X, Kuang TY, Xiang CB, Zhu JK, Oliver MJ, He YK (2015) The resurrection genome of Boea hygrometrica: a blueprint for survival of dehydration. Proc Natl Acad Sci U S A 112:5833–5837CrossRefGoogle Scholar
  26. Xu QL, Hu Z, Li CY, Wang XY, Wang CY (2009a) Tissue culture of Sinningia speciosa and analysis of the in vitro-generated tricussate whorled phyllotaxis (twp) variant. In Vitro Cell Dev Biol Plant 45:583–590CrossRefGoogle Scholar
  27. Xu WB, Pan B, Huang YS, Ye XX, Liu Y (2009b) Paraisometrum W. T. Wang, a newly recorded genus of Gesneriaceae from Guangxi, China. Guihaia 29:581–583 (In Chinese)Google Scholar
  28. Yang XY, Lü JF, da Silva JAT, Ma GH (2012) Somatic embryogenesis and shoot organogenesis from leaf explants of Primulina tabacum. Plant Cell Tissue Organ Cult 109:213–221CrossRefGoogle Scholar

Copyright information

© The Society for In Vitro Biology 2018

Authors and Affiliations

  1. 1.Germplasm Bank of Wild Species, Kunming Institute of BotanyChinese Academy of SciencesKunmingChina
  2. 2.College of Ecology and Soil & Water ConservationSouthwest Forestry UniversityKunmingChina
  3. 3.Key Laboratory of Economic Plants and Biotechnology, and Yunnan Key Laboratory for Wild Plant ResourceKunming Institute of Botany, Chinese Academy of SciencesKunmingChina
  4. 4.College of Life and Environmental SciencesMinzu University of ChinaBeijingChina

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