Abstract
Palicourea rigida, an endangered medicinal plant from the Brazilian Cerrado, produces the iridoid glucoside loganin as an active principle. The objectives of this study were to establish a micropropagation method for the species, to evaluate loganin accumulation in different genotypes cultivated in vitro, and to perform cDNA-sequence-related amplified polymorphism (cDNA-SRAP) analysis to discriminate the genotypes. Shoot cultures were initiated from apical explants derived from 7-day-old seedlings and incubated on agar medium without growth regulators. Nodal segments were excised from established shoots and cultured on agar medium supplemented with 6-benzylaminopurine (BAP), kinetin or 2-isopentenyl adenine. Rooting experiments in vitro were conducted using agar medium supplemented with 1-naphthaleneacetic acid or indole-3-butyric acid as well as common substrates (sand, sandy soil and clay soil), while rooting experiments ex vitro were performed using sandy soil or commercial Bioplant® substrate. Multiplication of shoots in vitro was improved by the presence of 0.1 µM BAP, while rooting only occurred in clay soil. Rooting under ex vitro conditions was improved when sandy soil was employed as substrate. High-performance liquid chromatographic analysis revealed that shoots cultured in the presence of 0.1 µM kinetin exhibited maximum levels of loganin (32.6 mg/g dry weight). cDNA-SRAP analysis was efficient in discriminating seven genotypes generated from seeds collected in the Cerrado. The genotypes were clustered into five groups, four of which had similar chemical profiles while the most genetically distant group accumulated no detectable amounts of loganin and could be classified as a chemotype.
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Abbreviations
- 2iP:
-
2-Isopentenyl adenine
- ANOVA:
-
Analysis of variance
- BAP:
-
6-Benzylaminopurine
- DEG:
-
Differentially expressed genes
- HPLC:
-
High-performance liquid chromatography
- IAA:
-
Indole-3-acetic acid
- IBA:
-
Indole-3-butyric acid (IBA)
- LOD:
-
Limit of detection
- LOQ:
-
Limit of quantification
- MS:
-
Murashige and Skoog
- NAA:
-
Naphthalene acetic acid (NAA)
- PCoA:
-
Principal coordinates analysis
- RT-PCR:
-
Reverse transcription-polymerase chain reaction
- S/N:
-
Signal to noise ratio
- SRAP:
-
Sequence-related amplified polymorphism
- TBE:
-
Tris–borate-EDTA buffer
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Acknowledgments
The authors wish to thank the Fundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP) for financial support for the study (Grant No. 07/58503-3).
Authors’ contribution
Study conception and design: AMSP, MV; Acquisition of data: MV, AMSP; Analysis and interpretation of data: MV, AMSP, BWB, SHTC; Drafting of manuscript: AMSP; Critical revision: BWB, SCF.
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Valdevite, M., Bertoni, B.W., Contini, S.H.T. et al. Accumulation of loganin by genotypes of Palicourea rigida and related differential gene expression as determined by cDNA-SRAP. Plant Cell Tiss Organ Cult 125, 445–456 (2016). https://doi.org/10.1007/s11240-016-0959-8
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DOI: https://doi.org/10.1007/s11240-016-0959-8