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Morpho- and histodifferentiation of shoot regeneration of Billbergia zebrina (Helbert) Lindley nodular cultures

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Abstract

Regeneration of nodular cultures involves the production of new individuals from organogenic nodules. Despite the existence of well-established protocols for bromeliads species, many gaps in understanding the mechanisms of this route still remain, particularly the regeneration of new shoots, a stage little explored. Therefore, this work aimed to monitor the morpho and histodifferentiation processes of shoot regeneration from nodular cultures of Billbergia zebrina. To accomplish this, nodular cultures previously obtained from nodal segments of B. zebrina were transferred to regeneration induction medium consisting of semi-solid MS medium supplemented with 1.0 µM of naphthalene acetic acid and 2.0 µM of benzylaminopurine, respectively. Cultures were maintained in a growth chamber over 8 weeks, and samples were collected weekly for analysis under light, transmission electron, and scanning electron microscopy. Cells in the shoot formation region presented characteristics of mitotically active cells, such as the presence of numerous plasmodesmata and conspicuous microtubules. New shoots maintained vascular connection to the node. Cells binding the nodule vascular bundle toward the shoot showed degeneration of organelles, which, in turn, originated vascular neoelements. The dome presented a depressed or prominent surface, depending on shoot development. Rounded stomata were conspicuous in the leaf primordia and generally higher in number as compared to the epidermal surface. This study elucidated the nodular culture in vitro system in aspects associated to the regeneration of shoots and it is anticipated that such data will lead to the optimization of present nodular cultures based protocols with the aim of conserving endangered bromeliad species.

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Abbreviations

AAN:

Naphthalene acetic acid

BAP:

Benzylaminopurine

BD:

Bud

CC:

Companion cell

CL:

Chlorenchyma

CP:

Chloroplast

CO:

Corpus

DM:

Donut-shaped mitochondria

DO:

Dome

ER:

Endoplasmic reticulum

GC:

Golgi complex

HY:

Hydrenchyma

LAVEG:

Plant Anatomy Laboratory

LCME:

Central Laboratory of Electron Microscopy

LF:

Leaf

LFDGV:

Laboratory of Plant Development and Genetics

LP:

Leaf primordia

LM:

Light microscopy

MI:

Mitochondria

MS:

Mitochondria in series

NC:

Nodular culture

NO:

Nodule

PL:

Plastoglobuli

RO:

Root

SEM:

Scanning electron microscopy

SH:

Shoot

SN:

Sieve neolelement

ST:

Stoma

TDZ:

Thidiazuron

TEM:

Transmission electron microscopy

TU:

Tunica

VC:

Vascularization

VA:

Vacuole

VB:

Vascular bundle

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Acknowledgments

This work was supported by the National Council for Scientific and Technological Development (CNPq, Brazil), Coordination for the Improvement of Higher Education Personnel (CAPES, Brazil), and FAPESC (Proc. 2780/2012-4). The authors acknowledge the staff of the Central Laboratory of Electron Microscopy (LCME), Plant Anatomy Laboratory (LAVEG) and Physiology Laboratory of Plant Development and Genetics (LFDGV) of the Federal University of Santa Catarina, Brazil.

Author contributions

TVS planned and designed all experiments, performed in vitro cultures, performed anatomical studies, collected and analyzed data, wrote manuscript, prepared figures; JFT maintained the cultures of plant material, collected data, contributed to manuscript writing; MPG performed analyzes of in vitro culture and anatomical studies, and contributed to manuscript writing; MS contributed in the planning of the experiments, performed anatomical studies, analyzed data and contributed to manuscript writing.

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Authors and Affiliations

  1. Programa de Pós-Graduação em Recursos Genéticos Vegetais, Universidade Federal de Santa Catarina, Rod. Admar Gonzaga, Km 3, Florianópolis, SC, 88034-001, Brazil

    Thaysi Ventura de Souza, Miguel Pedro Guerra & Marisa Santos

  2. Departamento de Botânica, Universidade Federal de Santa Catarina, Campus Universitário, Florianópolis, SC, 88040-900, Brazil

    Julia Faillace Thiesen & Marisa Santos

  3. Instituto Federal de Educação, Ciência e Tecnologia Catarinense, Campus Camboriú, Rua Joaquim Garcia, sn, Camboriú, SC, 88340-055, Brazil

    Thaysi Ventura de Souza

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  1. Thaysi Ventura de Souza
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  2. Julia Faillace Thiesen
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  3. Miguel Pedro Guerra
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  4. Marisa Santos
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Correspondence to Thaysi Ventura de Souza.

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de Souza, T.V., Thiesen, J.F., Guerra, M.P. et al. Morpho- and histodifferentiation of shoot regeneration of Billbergia zebrina (Helbert) Lindley nodular cultures. Plant Cell Tiss Organ Cult 127, 393–403 (2016). https://doi.org/10.1007/s11240-016-1061-y

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