In planta localisation patterns of MADS domain proteins during floral development in Arabidopsis thaliana

Background: MADS domain transcription factors play important roles in various developmental processes in flowering plants. Members of this family play a prominent role in the transition to flowering and the specification of floral organ identity. Several studies reported mRNA expression patterns of the genes encoding these MADS domain proteins, however, these studies do not provide the necessary information on the temporal and spatial localisation of the proteins. We have made GREEN FLUORESCENT PROTEIN (GFP) translational fusions with the four MADS domain proteins SEPALLATA3, AGAMOUS, FRUITFULL and APETALA1 from the model plant Arabidopsis thaliana and analysed the protein localisation patterns in living plant tissues by confocal laser scanning microscopy (CLSM). Results: We unravelled the protein localisation patterns of the four MADS domain proteins at a cellular and subcellular level in inflorescence and floral meristems, during development of the early flower bud stages, and during further differentiation of the floral organs. The protein localisation patterns revealed a few deviations from known mRNA expression patterns, suggesting a non-cell autonomous action of these factors or alternative control mechanisms. In addition, we observed a change in the subcellular localisation of SEPALLATA3 from a predominantly nuclear localisation to a more cytoplasmic localisation, occurring specifically during petal and stamen development. Furthermore, we show that the down-regulation of the homeodomain transcription factor WUSCHEL in ovular tissues is preceded by the occurrence of both AGAMOUS and SEPALLATA3 proteins, supporting the hypothesis that both proteins together suppress WUSCHEL expression in the ovule. Conclusion: This approach provides a highly detailed in situ map of MADS domain protein presence during early and later stages of floral development. The subcellular localisation of the transcription factors in the cytoplasm, as observed at certain stages during development, points to mechanisms other than transcriptional control. Together this information is essential to understand the role of these proteins in the regulatory processes that drive floral development and leads to new hypotheses

Comparative analysis of the complete plastomes of garlic Allium sativum and bulb onion Allium cepa

Sequencing and comparative characterization of plant plastid genomes, or plastomes, is an important tool for modern phylogenetic and taxonomic studies, as well as for understanding the plastome evolution. The genus Allium L. (family Amaryllidaceae) incorporates more than 900 species, includes economically signifi­cant vegetable crops such as garlic A. sativum, onion A. cepa, leek A. porrum, etc. In this work, the plastome of garlic A. sativum has been completely sequenced. The A. sativum plastome is 153172 bp in size. It consists of a large unique (LSC, 82035 bp) and small unique (SSC, 18015 bp) copies, separated by inverted repeats (IRa and IRb) of 26561 bp each. In the garlic plastome, 134 genes have been annotated: 82 protein-coding genes, 38 tRNA genes, 8 rRNA genes, and 6 pseudogenes. Comparative analysis of A. sativum and A. cepa plastomes reveals differences in the sizes of structural elements and spacers at the inverted repeat bound­aries. The total numbers of genes in A. sativum and A. cepa are the same, but the gene composition is dif­ferent: the rpl22 gene is functional in A. sativum, being a pseudogene in A. cepa; conversely, the rps16 gene is a pseudogene in A. sativum and a protein-coding gene in A. cepa. In the A. sativum and A. cepa plastomes, 32 SSR sequences have been identified. More than half of them are dinucleotides, and the remaining are tetra-, penta-, and hexanucleotides at the same time, trinucleotides were absent. The compared plastomes differ in the numbers of certain SSRs, and some are present in only one of the species

5′-UTR allelic variants and expression of the lycopene-ɛ-cyclase <i>LCYE</i> gene in maize (<i>Zea mays</i> L.) inbred lines of Russian selection

In breeding, biofortification is aimed at enriching the edible parts of the plant with micronutrients. Within the framework of this strategy, molecular screening of collections of various crops makes it possible to determine allelic variants of genes, new alleles, and the linkage of allelic variants with morphophysiological traits. The maize (Zea mays L.) is an important cereal and silage crop, as well as a source of the main precursor of vitamin A – β-carotene, a derivative of the β,β-branch of the carotenoid biosynthesis pathway. The parallel β,ε-branch is triggered by lycopene-ε-cyclase LCYE, a low expression of which leads to an increase in provitamin A content and is associated with the variability    of the 5’-UTR gene regulatory sequence. In this study, we screened a collection of 165 maize inbred lines of Russian selection for 5’-UTR LCYE allelic variants, as well as searched for the dependence of LCYE expression levels on the 5’-UTR allelic variant in the leaves of 14 collection lines. 165 lines analyzed were divided into three groups carrying alleles A2 (64 lines), A5 (31) and A6 (70), respectively. Compared to A2, allele A5 contained two deletions (at positions -267–260 and -296–290 from the ATG codon) and a G251→T substitution, while allele A6 contained one deletion (-290–296) and two SNPs (G251→T, G265→T). Analysis of LCYE expression in the leaf tissue of seedlings from accessions of 14 lines differing in allelic variants showed no associations of the 5’-UTR LCYE allele type with the level of gene expression. Four lines carrying alleles A2 (6178-1, 6709-2, 2289-3) and A5 (5677) had a significantly higher level of LCYE gene expression (~0.018–0.037) than the other 10 analyzed lines (~0.0001–0.004), among which all three allelic variants were present

Genome variability of domestic tomato varieties: data from AFLP analysis

Tomato Solanum lycopersicum L. is one of the main vegetable crops, accessions and cultivars of which are characterized by a low level of genomic polymorphism. Introgressive tomato breeding uses related wild Solanum species to improve cultivars for stress tolerance and fruit quality traits. The aim of this work was to evaluate the genome variability of 59 cultivars and perspective breeding lines of S. lycopersicum and 11 wild tomato species using the AFLP method. According to the AFLP analysis, four combinations of primers E32/M59, E32/M57, E38/M57, and E41/M59, which had the highest PIC (polymorphism information content) values, were selected. In the process of genotyping a collection of 59 cultivars/lines of S. lycopersicum and 11 wild tomato accessions, the selected primers revealed 391 fragments ranging in size from 80 to 450 bp, of which 114 fragments turned out to be polymorphic and 25 were unique. Analysis of the amplification spectra placed wild tomato accessions into separate clades. Sister clades included cultivars of FSCV breeding resistant to drought and/or cold and, in part, to late blight, Alternaria, Septoria, tobacco mosaic virus and blossom end rot, as well as tomato accessions not characterized according to these traits, which suggests that they have resistance to stress factors. In accessions of distant clades, there was clustering on the basis of resistance to Verticillium, cladosporiosis, Fusarium, tobacco mosaic virus, gray rot, and blossom end rot. The combination of accessions according to their origin from the originating organization was shown. The primer combinations E32/M59, E32/M57, E38/M57 and E41/M59 were shown to be perspective for genotyping tomato cultivars to select donors of resistance to various stress factors. The clade-specific fragments identified in this work can become the basis for the development of AFLP markers for traits of resistance to stress factors

Structural and functional features of phytoene synthase isoforms PSY1 and PSY2 in pepper Capsicum annuum L. cultivars

The fruits of various pepper cultivars are characterized by a different colour, which is determined by the pigment ratio; carotenoids dominate in ripe fruits, while chlorophylls, in immature fruits. A key regulator of carotenoid biosynthesis is the phytoene synthase encoded by the PSY gene. The Capsicum annuum genome contains two isoforms of this enzyme, localized in leaf (PSY2) and fruit (PSY1) plastids. In this work, the complete PSY1 and PSY2 genes were identified in nine C. annuum cultivars, which differ in ripe fruit colour. PSY1 and PSY2 sequence variability was 2.43 % (69 SNPs) and 1.21 % (36 SNPs). The most variable were PSY1 proteins of the cultivars ‘Maria’ (red-fruited) and ‘Sladkij shokolad’ (red-brown-fruited). All identified PSY1 and PSY2 homologs contained the phytoene synthase domain HH-IPPS and the transit peptide. In the PSY1 and PSY2 HH-IPPS domains, functionally significant sites were determined. For all accessions studied, the active sites (YAKTF and RAYV), aspartate-rich substrate-Mg2+-binding sites (DELVD and DVGED), and other functional residues were shown to be conserved. Transit peptides were more variable, and their similarity in the PSY1 and PSY2 proteins did not exceed 78.68 %. According to the biochemical data obtained, the largest amounts of chlorophylls and carotenoids across the cultivars studied were detected in immature and ripe fruits of the cv. ‘Sladkij shokolad’ and ‘Shokoladnyj’. Also, ripe fruits of the cv. ‘Nesozrevayuschij’ (green-fruited) were marked by significant chlorophyll content, but a minimum of carotenoids. The PSY1 and PSY2 expression patterns were determined in the fruit pericarp at three ripening stages in ‘Zheltyj buket’, ‘Sladkij shokolad’, ‘Karmin’ and ‘Nesozrevayuschij’, which have different ripe fruit colours: yellow, red-brown, dark red and green, respectively. In the leaves of the cultivars studied, PSY1 expression levels varied significantly. All cultivars were characterized by increased PSY1 transcription as the fruit ripened; the maximum transcription level was found in the ripe fruit of ‘Sladkij shokolad’, and the lowest, in ‘Nesozrevayuschij’. PSY2 transcripts were detected not only in the leaves and immature fruits, but also in ripe fruits. Assessment of a possible correlation of PSY1 and PSY2 transcription with carotenoid and chlorophyll content revealed a direct relationship between PSY1 expression level and carotenoid pigmentation during fruit ripening. It has been suggested that the absence of a typical pericarp pigmentation pattern in ‘Nesozrevayuschij’ may be associated with impaired chromoplast formation

INFLUENCE OF A CULTIVAR AND CONDITIONS OF VEGETATION ON SOWING FEATURES OF BARLEY DURING STORAGE

The experiments revealed the effect of the year of spring barley cultivation upon preservation of laboratory germination during storage. The seeds of spring barley varieties ‘Novichok’, ‘Lel’, ‘Tandem’ cultivated under contrasting vegetation conditions in Kirov region in 2007-08 have been laid for storage after the first year yield. The studies showed specificity of the barley cultivar according to the ability to preserve seed germination during long storage. The cultivar ‘Lel’ is characterized by the largest potential of biological longevity, which kept seed germination during five years of storage. The varieties differed in the time of seed aging. During all period of storage, laboratory germination of the varieties ‘Lel’, ‘Novichok’ and ‘Tandem’ was reduced on 3,0%, 3,8% and 6,5% respectively. The analysis of weather conditions was carried out during the period of grain ripening. The results of double factor variance analysis showed a significant effect of the factors ‘variety’, ‘year’ and their interaction on safety of laboratory seed germination during long storage. There are some authentic distinctions in germination among the seeds, grown in the years with different growing conditions. The studies showed that barley cultivars have various potential of agronomic longevity. The seeds of variety ‘Tandem’ kept their sowing properties during three years, the seeds of varieties ‘Lel’ and ‘Novichok’ kept their sowing properties during four-five years. It’s necessary to take into account the peculiarities of the cultivars and conditions of plant germination in the year of yield if the seeds are laid for a long storage