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Development of species-specific molecular markers in Vanilla for seedling selection of hybrids

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Abstract

Vanilla planifolia is the primary botanical source of vanilla extract used globally in various foods and beverages. V. planifolia has a global distribution based on a few foundational clones and therefore has limited genetic diversity. Many Vanilla species easily hybridize with V. planifolia and could be a source of valuable genetic traits like increased vanillin content, disease resistance, or early flowering. While breeding Vanilla hybrids may improve plant performance, basic molecular tools for this species are lacking. DNA-based molecular markers are the most efficient method to validate hybrid progeny, detect hybrids in commercial plantings, and identify unknown accessions. This study used publicly available sequence data to develop species-specific, qRT-PCR-based molecular markers for Vanilla. Over 580,000 assembled sequence fragments were filtered for species specificity and twenty-two targets were selected for qRT-PCR screening. Ten targets differentially amplified among V. planifolia, V. pompona, V. phaeantha, and V. palmarum with ΔCT values as high as 17.58 between species. The ten targets were used to validate the parentage of hybrid progeny from controlled crosses with most hybrid progeny showing amplification patterns similar to both parents. The ten targets were also used to screen sixteen Vanilla species for specificity, and supported species assignments for unknown accessions including the detection of putative hybrids. This is the first report using species-specific, qRT-PCR-based molecular markers in Vanilla. These markers are inexpensive, simple to develop, and can rapidly screen large populations. These methods will enable the further development of species-specific molecular markers when creating Vanilla interspecific hybrid populations.

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Acknowledgements

This work was supported by a State Scholarship award from the China Scholarship Council to Dr. Juan Li; and by funds provided by the University of Florida Dean for Research to Dr. Alan Chambers. The authors also acknowledge Pamela Moon for her assistance with seed germination and Shelby Weber for her assistance with tissue culture of the hybrid populations.

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Author notes

  1. Juan Li

    Present address: Department of Horticulture, Zhongkai University of Agriculture and Engineering, 501 Zhongkai Rd., Guangzhou, 510225, Guangdong, China

  2. Juan Li and Lynhe Demesyeux have contributed equally to this work.

Authors and Affiliations

  1. Horticultural Sciences Department, Tropical Research and Education Center, 18905 SW 280th St., Homestead, FL, 33031, USA

    Juan Li, Lynhe Demesyeux, Maria Brym & Alan H. Chambers

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  1. Juan Li
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  2. Lynhe Demesyeux
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Contributions

JL, LD, and AC generated the qRT-PCR data. MB maintained the tissue culture materials and performed the DNA extractions. AC designed the study, conducted the bioinformatics analysis, and wrote the first draft of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Alan H. Chambers.

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11033_2020_5287_MOESM1_ESM.xlsx

Supplementary file 1 Supplementary Table S1 Trinity assembly fragment ID, fragment DNA sequence, fragment length, and the number of mapped reads for each publicly available vanilla accession with GBS data. (XLSX 22 kb)

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Li, J., Demesyeux, L., Brym, M. et al. Development of species-specific molecular markers in Vanilla for seedling selection of hybrids. Mol Biol Rep 47, 1905–1920 (2020). https://doi.org/10.1007/s11033-020-05287-9

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