β-glucans, SAM, and GSH fluctuations in barley anther tissue culture conditions affect regenerants' DNA methylation and GPRE.

Background: Microspore embryogenesis is a process that produces doubled haploids in tissue culture environments and is widely used in cereal plants. The efficient production of green regenerants requires stresses that could be sensed at the level of glycolysis, followed by the Krebs cycle and electron transfer chain. The latter can be affected by Cu(II) ion concentration in the induction media acting as cofactors of biochemical reactions, indirectly influencing the production of glutathione (GSH) and S-adenosyl-L-methionine (SAM) and thereby affecting epigenetic mechanisms involving DNA methylation (demethylation-DM, de novo methylation-DNM).

An insight into tissue culture-induced variation origin shared between anther culture-derived triticale regenerants.

Background: The development of the plant in vitro techniques has brought about the variation identified in regenerants known as somaclonal or tissue culture-induced variation (TCIV). S-adenosyl-L-methionine (SAM), glutathione (GSH), low methylated pectins (LMP), and Cu(II) ions may be implicated in green plant regeneration efficiency (GPRE) and TCIV, according to studies in barley (Hordeum vulgare L.) and partially in triticale (× Triticosecale spp.

Polystyrene nanoparticles induce concerted response of plant defense mechanisms in plant cells.

Recent advances in knowledge suggest that micro- and nanoplastics pose a threat to plant health, however, the responses of plants to this stressor are not well-known. Here we examined the response of plant cell defence mechanisms to nanoparticles of commonly used plastic, polystyrene. We used plant cell cultures of widely cultivated plants, the monocots wheat and barley (Triticum aestivum L.

Evaluation of CRISPR/Cas9 Constructs in Wheat Cell Suspension Cultures.

Despite intensive optimization efforts, developing an efficient sequence-specific CRISPR/Cas-mediated genome editing method remains a challenge, especially in polyploid cereal species such as wheat. Validating the efficacy of nuclease constructs prior to using them in planta is, thus, a major step of every editing experiment. Several construct evaluation strategies were proposed, with PEG-mediated plasmid transfection of seedling-derived protoplasts becoming the most popular.

Corrigendum: Glutathione and copper ions as critical factors of green plant regeneration efficiency of triticale anther culture.

[This corrects the article DOI: 10.3389/fpls.2022.

S-Adenosyl-L-Methionine and Cu(II) Impact Green Plant Regeneration Efficiency.

The biological improvement of triticale, a cereal of increasing importance in agriculture, may be accelerated via the production of doubled haploid lines using in vitro culture. Among the relevant factors affecting the culture efficiency are Cu(II) or Ag(I) acting, e.g.

Glutathione and copper ions as critical factors of green plant regeneration efficiency of triticale anther culture.

Plant tissue culture techniques are handy tools for obtaining unique plant materials that are difficult to propagate or important for agriculture. Homozygous materials derived through cultures are invaluable and significantly accelerate the evaluation of new varieties, e.g.

Functional Validation of /guideRNA Constructs for Site-Directed Mutagenesis of Triticale .

Cas endonuclease-mediated genome editing provides a long-awaited molecular biological approach to the modification of predefined genomic target sequences in living organisms. Although /guide (g)RNA constructs are straightforward to assemble and can be customized to target virtually any site in the plant genome, the implementation of this technology can be cumbersome, especially in species like triticale that are difficult to transform, for which only limited genome information is available and/or which carry comparatively large genomes. To cope with these challenges, we have pre-validated /gRNA constructs (1) by frameshift restitution of a reporter gene co-introduced by ballistic DNA transfer to barley epidermis cells, and (2) via transfection in triticale protoplasts followed by either a T7E1-based cleavage assay or by deep-sequencing of target-specific PCR amplicons.

Copper Ions Induce DNA Sequence Variation in Zygotic Embryo Culture-Derived Barley Regenerants.

tissue culture could be exploited to study cellular mechanisms that induce sequence variation. Altering the metal ion composition of tissue culture medium affects biochemical pathways involved in tissue culture-induced variation. Copper ions are involved in the mitochondrial respiratory chain and Yang cycle.

Heritability of meiotic restitution and fertility restoration in haploid triticale.

A single division meiosis mechanism of meiotic restitution is incompletely penetrant but significantly associated with restored fertility in triticale haploids (n = 21, genome formula ABR). Meiotic restitution, or failure of meiosis to produce gametes with a reduced chromosome number, can lead to the restoration of fertility in allohaploids. Meiotic restitution is of major interest for producing doubled haploids, as haploid plants undergoing meiotic restitution can often form seeds without the need to apply mitosis inhibitors to double chromosome number.

The Effects of Combined Abiotic and Pathogen Stress in Plants: Insights From Salinity and pv Interaction in Cucumber.

Plants are often challenged by abiotic and biotic stresses acting in combination and the response to combinatorial stress differs from that triggered by each factor individually. Although salinity and pathogens are major stressors limiting plant growth and productivity worldwide, their interaction is poorly understood. The reactions to pathogens overlap with those to abiotic stresses, and reactive oxygen species (ROS) and stress hormones represent central nodes in the interacting signaling pathways.

DNA methylation changes and TE activity induced in tissue cultures of barley (Hordeum vulgare L.).

Background: In vitro plant regeneration via androgenesis or somatic embryogenesis is capable of inducing (epi)mutations that may affect sexual progenies. While epimutations are associated with DNA methylation, mutations could be due to the movement of transposons. The common notion is that both processes are linked.

Tissue culture-induced genetic and epigenetic variation in triticale (× Triticosecale spp. Wittmack ex A. Camus 1927) regenerants.

Plant regeneration via in vitro culture can induce genetic and epigenetic variation; however, the extent of such changes in triticale is not yet understood. In the present study, metAFLP, a variation of methylation-sensitive amplified fragment length polymorphism analysis, was used to investigate tissue culture-induced variation in triticale regenerants derived from four distinct genotypes using androgenesis and somatic embryogenesis. The metAFLP technique enabled identification of both sequence and DNA methylation pattern changes in a single experiment.

Extended metAFLP approach in studies of tissue culture induced variation (TCIV) in triticale.

We present the development of the theoretical background of the metAFLP approach which allows for partition of complex variation into sequence changes, de novo methylation and demethylation of the regenerants derived via in vitro tissue culture methods in the case of triticale. It was demonstrated that, independent of whether andro- or embryogenesis was used for plant regeneration, the level of sequence changes identified between regenerants is about 10 %. Moreover, DNA demethylation prevails over de novo methylation of the regenerants compared to the donor plant.

Analysis of T-DNA integration and generative segregation in transgenic winter triticale (x Triticosecale Wittmack).

Background: While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species.

Results: Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT) and a synthetic green fluorescent protein gene (gfp).

Aneuploidy among androgenic progeny of hexaploid triticale (XTriticosecale Wittmack).

Doubled haploids are an established tool in plant breeding and research. Of several methods for their production, androgenesis is technically simple and can efficiently produce substantial numbers of lines. It is well suited to such crops as hexaploid triticale.

Quantification of the tissue-culture induced variation in barley (Hordeum vulgare L.).

Background: When plant tissue is passaged through in vitro culture, many regenerated plants appear to be no longer clonal copies of their donor genotype. Among the factors that affect this so-called tissue culture induced variation are explant genotype, explant tissue origin, medium composition, and the length of time in culture. Variation is understood to be generated via a combination of genetic and/or epigenetic changes.

The application of the AFLP method to determine the purity of homozygous lines of barley (Horedum vulgare L.).

Amplified fragment length polymorphism of DNA has been used to analyse the equality of plants obtained from isolated microspores. Although the control parental material was regarded as being highly homozygous, the analysis of the banding patterns of single plants showed a certain level of polymorphism. The analysis of regenerants with a doubled chromosome number did not show any diversity within the progeny of a single line.