Androgenic haploid plant development via embryogenesis with simultaneous determination of bioactive metabolites in Cambod tea (Camellia assamica ssp. lasiocalyx)

Vijay Kumar Mishra,Ruchira Bajpai,Rakhi Chaturvedi

doi:10.1007/s11240-021-02203-2

Vijay Kumar Mishra, Ruchira Bajpai + Show 1 more

Open Access

https://doi.org/10.1007/s11240-021-02203-2

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Abstract

This pioneering work reports successful androgenic plant development via embryogenesis from microspore calluses in anther cultures and estimation of bioactive metabolites in in vitro regenerants and parent plant (control) of Cambod tea, Camellia assamica ssp. lasiocalyx (Planch MS) cultivar TV19. Anthers bearing microspores at early-to-late uni-nucleate stage were selected to initiate androgenesis. A pre-treatment of 5 °C for 5 days in the dark was most effective to initiate profusely growing white callusing from microspores within 10 weeks of culture on MS medium (6% sucrose) supplemented with high cytokinin/auxin ratio maintained by 6-benzylaminopurine (BAP) and 2,4-dichlorophenxoyacetic acid. Nodular structures on the callus surface differentiated into embryos. Further developement of the embryos occurred on embryogenesis medium but, with ten times reduced concentration of growth regulators and additives. Germination of embryos into complete plantlets was achieved when major salts in medium were reduced to half MS (½ MS) and augmented with BAP, GA3 and IBA along with glutamine and serine. Cytological examination of the root-tip cells revealed that regenerated plantlets were haploids (2n = x = 15), which was further confirmed through flow cytometry. The hot-water extracts from in vitro haploid calluses, embryos and field-grown donor plant were utilized for quantification of (+)-catechin, (−)-epicatechin, (−)-epigallocatechin gallate, caffeine and theophylline. Our findings revealed that the metabolite profile of in vitro regenerated haploid cultures is comparable to that of the mother plant, thereby presenting them as potential source for genome duplication and development of genetically stable homozygous pure breeding lines. This is first report on haploids in out-breeding tree, Cambod tea. It’s a significant achievement towards generating homozygous lines, which is impossible using conventional methods. Haploids showed consistent metabolite production.

Highlights

Camellia assamica ssp. lasiocalyx (Planch mass spectrometry (MS)) or Cambod tea belongs to the Theaceae family (Parmar et al 2012; Meegahakumbura et al 2018)
Our findings revealed that the metabolite profile of in vitro regenerated haploid cultures is comparable to that of the mother plant, thereby presenting them as potential source for genome duplication and development of genetically stable homozygous pure breeding lines
The therapeutic potential and aromatic characteristics of this commercially valuable crop is mainly attributed to the presence of polyphenol catechins including (+)- catechin (C), (-)-epicatechin (EC), (-)-gallocatechin (GC), (-)epigallocatechin (EGC), (- )-catechin gallate (CG), (-)-gallocatechin gallate (GCG), (-)-epicatechin gallate (ECG) and (-)epigallocatechin gallate (EGCG) and purine alkaloids, such as caffeine, theobromine, theophylline and theacrine (Wei et al 2011; Khan and Mukhtar 2019)

Summary

Introduction

Camellia assamica ssp. lasiocalyx (Planch MS) or Cambod tea belongs to the Theaceae family (Parmar et al 2012; Meegahakumbura et al 2018). Lasiocalyx (Planch MS) or Cambod tea belongs to the Theaceae family (Parmar et al 2012; Meegahakumbura et al 2018) It is an evergreen perennial tree, and exhibits high cross pollinating behavior (Olaniyi et al 2014; Xia et al 2020). Polyphenols (TP) are a class of compounds with free radical scavenging property that monitor the action of various oxidases found in the body (Yan et al 2020). This characteristic of polyphenols is due to the phenolic hydroxyl structure, where the electrons exhibit conjugation effect that weakens the binding ability of the hydrogen making it more susceptible to dissociation. The above listed biotic and abiotic elements in combination with the highly variable global environment lead to inconsistent metabolite production reducing the flavor quality of this valuable cash crop (Ahmed et al 2019)

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