Genome-wide analysis of the C2H2-ZFP gene family in Stevia rebaudiana reveals involvement in abiotic stress response.

Stevia (Stevia rebaudiana Bertoni) is a natural sweetener plant that accumulates highly sweet steviol glycosides (SGs) especially in leaves. Stevia is native to humid areas and does not have a high tolerance to drought which is the most serious abiotic stress restricting its production worldwide. C2H2 zinc finger proteins (C2H2-ZFPs) are a group of well-known transcription factors that involves in various developmental, physiological and biochemical activities as well as in response to abiotic stresses. Here we analyzed C2H2-ZFP gene family in stevia and identified a total of 185 putative SrC2H2-ZF proteins from the genome sequence of S. rebaudiana. We further characterized the identified C2H2-ZF domains and their organization, additional domains and motifs and analyzed their physicochemical properties, localization and gene expression patterns. The cis-element analysis suggested multiple roles of SrC2H2-ZFPs in response to light, phytohormone, and abiotic stresses. In silico analysis revealed that the stevia C2H2-ZFP genes are interactively expressed in different tissues and developmental stages and some C2H2-ZFP genes are involved in response to drought stress. This study provides a background for future exploration of the functional, and regulatory aspects of the C2H2-ZFP gene family in S. rebaudiana.

Meiotic segregation and post-meiotic drive of the Festuca pratensis B chromosome.

In many species, the transmission of B chromosomes (Bs) does not follow the Mendelian laws of equal segregation and independent assortment. This deviation results in transmission rates of Bs higher than 0.5, a process known as "chromosome drive". Here, we studied the behavior of the 103 Mbp-large B chromosome of Festuca pratensis during all meiotic and mitotic stages of microsporogenesis. Mostly, the B chromosome of F. pratensis segregates during meiosis like standard A chromosomes (As). In some cases, the B passes through meiosis in a non-Mendelian segregation leading to their accumulation already in meiosis. However, a true drive of the B happens during the first pollen mitosis, by which the B preferentially migrates to the generative nucleus. During second pollen mitosis, B divides equally between the two sperms. Despite some differences in the frequency of drive between individuals with different numbers of Bs, at least 82% of drive was observed. Flow cytometry-based quantification of B-containing sperm nuclei agrees with the FISH data.

In vitro regeneration and Agrobacterium-mediated genetic transformation of Dragon's Head plant (Lallemantia iberica).

Dragon's head plant (Lallemantia iberica), is a flowering species belongs to the mint family (Lamiaceae). The species contains valuable essential oils, mucilage and oil which are used in pharmaceutical and food industries. Tissue culture is a feasible strategy to attain large-scale production of plantlets with a huge potential to produce plants with superior quality. The objective of this study was to develop a simple and efficient method for regeneration and transformation of L. iberica. To reach this goal, the regeneration ability of various explants including leaf, cotyledonary node, hypocotyl and cotyledon segments was investigated in MS medium supplemented with diverse concentrations of NAA (Naphthalene acetic acid) and BAP (6-Benzyl Amino Purine). According to the results, cotyledonary nodes showed the best regeneration response. The maximum rate of regeneration (and number of induced shoots was achieved in 1 mg l-1 BAP in combination with 0.05 mg l-1 NAA from the cotyledonary nodes. Additionally, through the optimized regeneration technique Agrobacterium-mediated transformation of L. iberica was successfully accomplished. Gene transfer was assessed on leaf samples from regenerated plantlets under a fluorescent microscope to detect the GFP signals. Moreover, transgene integration and its expression were confirmed by PCR and RT-PCR analysis, respectively. The establishment of these efficient regeneration and genetic transformation methods paved the way for further application such as plant improvement, functional analysis and gene editing.

Differentially Amplified Repetitive Sequences Among Aegilops tauschii Subspecies and Genotypes.

Genomic repetitive sequences commonly show species-specific sequence type, abundance, and distribution patterns, however, their intraspecific characteristics have been poorly described. We quantified the genomic repetitive sequences and performed single nucleotide polymorphism (SNP) analysis between 29 Ae. tauschii genotypes and subspecies using publicly available raw genomic Illumina sequence reads and used fluorescence in situ hybridization (FISH) to experimentally analyze some repeats. The majority of the identified repetitive sequences had similar contents and proportions between anathera, meyeri, and strangulata subspecies. However, two Ty3/gypsy retrotransposons (CL62 and CL87) showed significantly higher abundances, and CL1, CL119, CL213, CL217 tandem repeats, and CL142 retrotransposon (Ty1/copia type) showed significantly lower abundances in subspecies strangulata compared with the subspecies anathera and meyeri. One tandem repeat and 45S ribosomal DNA (45S rDNA) abundances showed a high variation between genotypes but their abundances were not subspecies specific. Phylogenetic analysis using the repeat abundances of the aforementioned clusters placed the strangulata subsp. in a distinct clade but could not discriminate anathera and meyeri. A near complete differentiation of anathera and strangulata subspecies was observed using SNP analysis; however, var. meyeri showed higher genetic diversity. FISH using major tandem repeats couldn't detect differences between subspecies, although (GAA)10 signal patterns generated two different karyotype groups. Taken together, the different classes of repetitive DNA sequences have differentially accumulated between strangulata and the other two subspecies of Ae. tauschii that is generally in agreement with spike morphology, implying that factors affecting repeatome evolution are variable even among highly closely related lineages.

Production of synthetic wheat lines to exploit the genetic diversity of emmer wheat and D genome containing Aegilops species in wheat breeding.

Due to the accumulation of various useful traits over evolutionary time, emmer wheat (Triticum turgidum subsp. dicoccum and dicoccoides, 2n = 4x = 28; AABB), durum wheat (T. turgidum subsp. durum, 2n = 4x = 28; AABB), T. timopheevii (2n = 4x = 28; AAGG) and D genome containing Aegilops species offer excellent sources of novel variation for the improvement of bread wheat (T. aestivum L., AABBDD). Here, we made 192 different cross combinations between diverse genotypes of wheat and Aegilops species including emmer wheat × Ae. tauschii (2n = DD or DDDD), durum wheat × Ae. tauschii, T. timopheevii × Ae. tauschii, Ae. crassa × durum wheat, Ae. cylindrica × durum wheat and Ae. ventricosa × durum wheat in the field over three successive years. We successfully recovered 56 different synthetic hexaploid and octaploid F2 lines with AABBDD, AABBDDDD, AAGGDD, D1D1XcrXcrAABB, DcDcCcCcAABB and DvDvNvNvAABB genomes via in vitro rescue of F1 embryos and spontaneous production of F2 seeds on the Fl plants. Cytogenetic analysis of F2 lines showed that the produced synthetic wheat lines were generally promising stable amphiploids. Contribution of D genome bearing Aegilops and the less-investigated emmer wheat genotypes as parents in the crosses resulted in synthetic amphiploids which are a valuable resource for bread wheat breeding.

Repetitive DNA landscape in essential A and supernumerary B chromosomes of Festuca pratensis Huds.

Here, we characterized the basic properties of repetitive sequences in essential A and supernumerary B chromosomes of Festuca pratensis Huds. This was performed by comparative analysis of low-pass Illumina sequence reads of B chromosome lacking (-B) and B chromosome containing (+B) individuals of F. pratensis. 61% of the nuclear genome is composed of repetitive sequences. 43.1% of the genome are transposons of which DNA transposons and retrotransposons made up 2.3% and 40.8%, respectively. LTR retrotransposons are the most abundant mobile elements and contribute to 40.7% of the genome and divided into Ty3-gypsy and Ty1-copia super families with 32.97% and 7.78% of the genome, respectively. Eighteen different satellite repeats were identified making up 3.9% of the genome. Five satellite repeats were used as cytological markers for chromosome identification and genome analysis in the genus Festuca. Four satellite repeats were identified on B chromosomes among which Fp-Sat48 and Fp-Sat253 were specific to the B chromosome of F. pratensis.