Taxonomic significance of pollen morphology for species delimitation in <Emphasis Type="Italic">Psidium</Emphasis> (Myrtaceae)

Previous studies reported Psidium as one of the most difficult genera to delimit within the American Myrtaceae. Even though palynology has improved the taxonomy of Angiosperms, information about the usefulness of pollen morphology for taxonomic purposes in Myrtaceae remains contradictory. Here, we investigate the significance of pollen morphology for Psidium taxonomy with specific focus on its usefulness for determining species groups of taxonomic significance. Pollen traits observed by light and scanning electron microscopy were quantified and examined using cluster and ordination analyses. Average size of pollen grains was visualized by boxplots. Pollen grains of Psidium are isopolar, oblate, peroblate or oblate-spheroidal, 3-syncolporate or 4-syncolporate. The sexine ornamentation is rugulate, granulate or spinulose-granulate and differs between the mesocolpium and apocolpium. Cluster analysis revealed four distinct groups: Psidium cauliflorum (G1) and Psidium oligospermum (G3) as single-species groups; Psidium brownianum, P. oblongatum, P. ovale, P. sartorianum, P. guajava, Psidium sp. 1, Psidium sp. 2 (G2), and Psidium cattleianum, P. longipetiolatum, P. guineense, P. myrtoides (G4). Supported by ordination analysis, three traits better explained these groups: type of exine ornamentation, size of P-EV and pollen shape. The used approach efficiently distinguished related species, as well as explained species groups of taxonomic significance suggesting pollen morphology to be a significant source of information for taxonomic studies in Psidium.     

Development of an STS map of an interspecific progeny of <Emphasis Type="Italic">Malus</Emphasis>

Simple sequence repeat (SSR) markers developed from Malus, as well as Prunus, Pyrus and Sorbus, and some other sequence-tagged site (STS) loci were analysed in an interspecific F₁ apple progeny from the cross ‘Fiesta’ × ‘Totem’ that segregated for several agronomic characters. A linkage map was constructed using 259 STS loci (247 SSRs, four SCARs and eight known-function genes) and five genes for agronomic traits—scab resistance (Vf), mildew resistance (Pl-2), columnar growth habit (Co), red tissues (Rt) and green flesh background colour (Gfc). Ninety SSR loci and three genes (ETR1, Rt and Gfc) were mapped for the first time in apple. The transferability of markers from other Maloideae to Malus was found to be around 44%. The loci are spread across 17 linkage groups, corresponding to the basic chromosome number of Malus and cover 1,208 cM, approximately 85% of the estimated length of the apple genome. Interestingly, we have extended the top of LG15 with eight markers covering 25 cM. The average map density is 4.7 cM per marker; however, marker density varies greatly between linkage groups, from 2.5 in LG14 to 8.9 in LG7, with some areas of the genome still in need of further STS markers for saturation.     

A high-density linkage map with 2560 markers and its application for the localization of the male-sterile genes <Emphasis Type="Italic">ms3</Emphasis> and <Emphasis Type="Italic">ms4</Emphasis> in <Emphasis Type="Italic">Cryptomeria japonica</Emphasis> D. Don

Cryptomeria japonica pollinosis is one of the most serious allergic diseases in Japan; this is a social problem because C. japonica is the most important Japanese forestry species. In order to reduce the amount of pollen dispersed, breeding programs using trees with male-sterile genes have been implemented. High-density linkage maps with stable ordering of markers facilitate the localization of male-sterile genes and the construction of partial linkage maps around them in order to develop markers for use in marker-assisted selection. In this study, a high-density linkage map for C. japonica with 2560 markers was constructed. The observed map length was 1266.2 cM and the mean distance between adjacent markers was 0.49 cM. Using information from this high-density map, we newly located two male-sterile genes (ms3 and ms4) on the first and fourth linkage groups, respectively, and constructed partial linkage maps around these loci. We also constructed new partial linkage maps around the ms1 and ms2 loci using additional SNP markers. The closest markers to the ms1, ms2, ms3, and ms4 male-sterile loci were estSNP04188 (1.8 cM), estSNP00695 (7.0 cM), gSNP05415 (3.1 cM), and estSNP01408 (7.0 cM) respectively. These results allowed us to develop SNP markers tightly linked to the male sterile genes for use in MAS; this will accelerate the future isolation of these genes by map-based cloning approaches.     

Pollen ultrastructure in <Emphasis Type="Italic">Aristolochia manshuriensis</Emphasis> and <Emphasis Type="Italic">A. contorta</Emphasis> (Aristolochiaceae)

Pollen ultrastructure has been studied in two relict and rare species of the genus Aristolochia, A. contorta Bunge and A. manshuriensis Kom. (Aristolochiaceae). Both species have inaperturate, spheroidal, sometimes distally monocolpate or distally bicolpate pollen grains. The equatorial and polar axes of pollen grain in A. manshuriensis are 48.5 and 44.0 μm, respectively. The percentage of defective pollen grains in A. manshuriensis is 3.4%. The fossulate, perforated exine is up to 2.3 μm in thickness; the sexine and the nexine are almost equal in thickness. In A. contorta, the equatorial axis of pollen grain is 36.6 μm: the defectiveness percentage, 24.5%. The exine is verrucate, up to 0.3 μm in thickness, while the sexine is two to three times thicker than the nexine. The pollen germination experiments have shown that pollen of A. manshuriensis, in contrast to A. contorta, can germinate in 10–20% sucrose at 22°С. These data and the high percentage of pollen defectiveness in A. contorta indicate that the androecium function in this species is reduced. The reduction of the androecium function is evidenced by a small amount of pollen grains in anthers or empty anthers and a high percentage of defective pollen grains.     

Pollen morphology of selected species of the genera <Emphasis Type="Italic">Chrysodracon</Emphasis> and <Emphasis Type="Italic">Dracaena</Emphasis> (Asparagaceae,subfamily Nolinoideae) and its systematic implications

Pollen grains of 3 species from the genus Chrysodracon and 20 species from the genus Dracaena were examined by light, scanning electron and fluorescence microscopy. The basic shape of the pollen grains is subprolate and prolate, but prolate-spheroidal pollen grains may also be found. Pollen of Chrysodracon and Dracaeana is dispersed as monads, rarely as dyads. In terms of size, the pollen grains studied were classified as medium-sized and large. There are two different types of apertures: monosulcate and monoulcerate in Dracaena and monoulcerate in Chrysodracon. The surface of the non-apertural areas is psilate-perforate, irregularly folded, microreticulate and fossulate. Ornamentation of the apertural region is microreticulate, microreticulate-baculate, baculate, psilate-perforate, psilate-perforate-verrucate, granulate and irregularly folded. Irregular perforations are present, and the wall structure is tectate-columellate. Our results suggest that some species of Chrysodracon and Dracaena may be separated based on their pollen grain micromorphology on the distal region.     

Morphological variation in pollen grains of <Emphasis Type="Italic">Mucuna</Emphasis> (Leguminosae): new biogeographic and evolutionary patterns

Mucuna comprises 105 species with an overall pantropical distribution and is divided into three subgenera: M. subg. Mucuna, M. subg. Stizolobium and M. subg. Macrocarpa. Although phylogenetic studies have supported the occurrence of three main clades, evolutionary relationships among them are not fully resolved. The objective of this study was to examine pollen grain morphology from representatives of all three subgenera and map these onto the phylogenetic trees generated by analysis of other characters. Pollen grain surface, form, size, and aperture number were compared. A Bayesian inference tree using matK sequences was constructed. The results indicate that the representatives of M. subg. Macrocarpa have the smallest pollen grains in the genus (a synapomorphic character here identified for this subgenus) and that species of subgenus Mucuna (those with umbelliform inflorescences) have the largest pollen grains. Additional morphological diversity of the pollen grain surface was noted: reticulate and/or micro-reticulate (in all three subgenera), perforate, gemmate or verrucose (only in M. subg. Mucuna). For all studied taxa, the pollen grains are triaperturate, except for two species of M. subg. Mucuna, which have tetraperturate pollen. The phylogenetic tree obtained using the matK marker resolved M. subg. Stizolobium as the earliest diverging lineage in Mucuna. Based on this phylogeny, a reticulate ornamentation pattern of the pollen surface may represent the ancestral state for the genus, while the larger pollen size and the foraminate, gemmate, and verrucose ornamentations are derived characteristics within the genus. These putative derived ornamentations have been observed only in neotropical species.     

Orange color is associated with <Emphasis Type="Italic">CYC</Emphasis>-<Emphasis Type="Italic">B</Emphasis> expression in tomato fleshy fruit

Carotenes are plant secondary metabolites that are important for human health. Additionally, carotenes influence fruit color, which is a major trait for breeding. We compared the expression and sequences of genes related to color phenotypes in tomato inbred lines that produce different colors of fleshy fruit. Up-regulation of CYC-B expression and higher amount of β-carotene content in fruit ripening stage and nucleotide variations in the 5′ region of the gene were detected in orange fruited inbred lines compared to the other lines. Our results indicated that there is a close relationship between the expression pattern of the CYC-B gene and the orange color of fleshy fruit. We identified 4 SNPs in the promoter region of CYC-B genes associated with the orange fruit color. Moreover, the segregation ratio and color phenotypes in an F2 generation further indicated that one of the detected SNPs were associated with the orange color in the tested inbred lines. Our study provides valuable information to breeders for marker-assisted selection to produce desirable tomato varieties with health benefits by varying carotenoid levels.     

Morphological and genetic characterization of a new cytoplasmic male sterility system (<Emphasis Type="Italic">oxa</Emphasis> CMS) in stem mustard (<Emphasis Type="Italic">Brassica juncea</Emphasis>)

Key message

oxa CMS is a new cytoplasmic male sterility type in Brassica juncea.

Abstract

oxa CMS is a cytoplasmic male sterility (CMS) line that has been widely used in the production and cultivation of stem mustard in the southwestern China. In this study, different CMS-type specific mitochondrial markers were used to confirm that oxa CMS is distinct from the pol CMS, ogu CMS, nap CMS, hau CMS, tour CMS, Moricandia arvensis CMS, orf220-type CMS, etc., that have been previously reported in Brassica crops. Pollen grains of the oxa CMS line are sterile with a self-fertility rate of almost 0% and the sterility strain rate and sterility degree of oxa CMS is 100% due to a specific flower structure and flowering habit. Scanning electron microscopy revealed that most pollen grains in mature anthers of the oxa CMS line are empty, flat and deflated. Semi-thin section further showed that the abortive stage of anther development in oxa CMS is initiated at the late uninucleate stage. Abnormally vacuolated microspores caused male sterility in the oxa CMS line. This cytological study combined with marker-assisted selection showed that oxa CMS is a novel CMS type in stem mustard (Brassica juncea). Interestingly, the abortive stage of oxa CMS is later than those in other CMS types reported in Brassica crops, and there is no negative effect on the oxa CMS line growth period. This study demonstrated that this novel oxa CMS has a unique flower structure with sterile pollen grains at the late uninucleate stage. Our results may help to uncover the mechanism of oxa CMS in Brassica juncea.

     

Genetic Control of the Wavy Shoots Character in Radish <Emphasis Type="Italic">Raphanus sativus</Emphasis> L.

The results of genetic analysis of the wavy shoots character supported the hypothesis on the existence of two genes, Wsh1 and Wsh2, whose mutations caused wavy shoots in radish lines 16 and 26, respectively. As shown by analysis of joint inheritance with marker characters, wavy shoots were inherited independently of the biochemical markers (genes Lap1, Ep, Dia1, and Aat). Gene Wsh1 was shown to be linked to genes A, ac, and ar; the mode of inheritance depended on the cross combination. In hybrids with mutant line 16, plants with novel morphological defects, which were absent in the parental forms, were detected. Possible mechanisms of new morphological abnormalities and change of linkage in the combinations analyzed are discussed.     

Differentiation of the pollen size in five representatives of <Emphasis Type="Italic">Taraxacum</Emphasis> sect. <Emphasis Type="Italic">Palustria</Emphasis>

Measurements of the pollen size in 5 species of Taraxacum sect. Palustria at three levels of ploidy: 2n = 3x = 24 (T. paucilobum), 2n = 4x = 32 (T. vindobonense, T. trilobifolium), 2n = 5x = 40 (T. mendax) and one taxon of unknown number of chromosomes 2n = ? (T. portentosum) are presented in this paper. Obtained results indicate a lack of distinct positive correlation between the pollen size and ploidy in the studied group of plants. Distinct relationship was, however, found between ploidy and the range of pollen size and shape variability. Most variable were the pollen grains of triploid T. paucilobum and the least — those in pentaploid T. mendax. Ranges of pollen variability in tetraploid T. trilobifolium and T. vindobonense and in T. portentosum of unknown number of chromosomes showed intermediate values.     

<Emphasis Type="Italic">Cannabis</Emphasis> is indigenous to Europe and cultivation began during the Copper or Bronze age: a probabilistic synthesis of fossil pollen studies

Conventional wisdom states Cannabis sativa originated in Asia and its dispersal to Europe depended upon human transport. Various Neolithic or Bronze age groups have been named as pioneer cultivators. These theses were tested by examining fossil pollen studies (FPSs), obtained from the European Pollen Database. Many FPSs report Cannabis or Humulus (C/H) with collective names (e.g. Cannabis/Humulus or Cannabaceae). To dissect these aggregate data, we used ecological proxies to differentiate C/H pollen, as follows: unknown C/H pollen that appeared in a pollen assemblage suggestive of steppe (Poaceae, Artemisia, Chenopodiaceae) we interpreted as wild-type Cannabis. C/H pollen in a mesophytic forest assemblage (Alnus, Salix, Populus) we interpreted as Humulus. C/H pollen curves that upsurged and appeared de novo alongside crop pollen grains we interpreted as cultivated hemp. FPSs were mapped and compared to the territories of archaeological cultures. We analysed 479 FPSs from the Holocene/Late Glacial, plus 36 FPSs from older strata. The results showed C/H pollen consistent with wild-type C. sativa in steppe and dry tundra landscapes throughout Europe during the early Holocene, Late Glacial, and previous glaciations. During the warm and wet Holocene Climactic Optimum, forests replaced steppe, and Humulus dominated. Cannabis retreated to steppe refugia. C/H pollen consistent with cultivated hemp first appeared in the Pontic-Caspian steppe refugium. GIS mapping linked cultivation with the Copper age Varna/Gumelni?a culture, and the Bronze age Yamnaya and Terramara cultures. An Iron age steppe culture, the Scythians, likely introduced hemp cultivation to Celtic and Proto-Slavic cultures.     

Morphology and variability of pollen of the genus <Emphasis Type="Italic">Spiraea</Emphasis> L. (Rosaceae) in Siberia and the Far East

Pollen of 12 species of the genus Spirea L. growing in different regions of Siberia and the Far East was investigated. The ultrasculpture of pollen grains is described. On the basis of aperture morphology and exine sculpture, pollen grains are typified, with key indices given for their identification. The species of the Spiraria section are distinguished by large grains. Pollen grains of S. media and S. alpina considerably vary in size. The level of natural polymorphism of palynological indices has been established.     

Detection of Insertions/Deletions Within <Emphasis Type="Italic">SIRT1</Emphasis>, <Emphasis Type="Italic">SIRT2</Emphasis> and <Emphasis Type="Italic">SIRT3</Emphasis> Genes and Their Associations with Body Measurement Traits in Cattle

Growth traits are complex quantitative traits controlled by numerous candidate genes, and they can be well-evaluated using body measurement traits. As the members of the nicotinamide adenine dinucleotide-dependent family of histone deacetylases, class I sirtuin genes (including SIRT1, SIRT2 and SIRT3) play crucial roles in regulating lipid metabolism, cellular growth and metabolism, suggesting that they are potential candidate genes affecting body measurement traits in animals. Hence, the objective of this work aimed to detect novel insertions/deletions (indels) of SIRT1, SIRT2 and SIRT3 genes in 955 cattle belonging to five breeds, as well as to evaluate their effects on body measurement traits. Herein, the novel 12-bp indel of SIRT1 gene, the 7-bp indel of SIRT2 gene and the 26-bp indel of SIRT3 gene were firstly reported, respectively. The association analysis indicated that the indels within SIRT1 and SIRT2 genes were significantly associated with body measurement traits such as body weight, chest circumference, height at hip cross, hip width, body height, etc. (P?<?0.05 or P?<?0.01). Therefore, based on these findings, the two novel indel variants within bovine SIRT1 and SIRT2 genes could be considered as potential molecular markers for growth traits in cattle selection practices and breeding.     

Proline synthesis in developing microspores is required for pollen development and fertility

Background

In many plants, the amino acid proline is strongly accumulated in pollen and disruption of proline synthesis caused abortion of microspore development in Arabidopsis. So far, it was unclear whether local biosynthesis or transport of proline determines the success of fertile pollen development.

Results

We analyzed the expression pattern of the proline biosynthetic genes PYRROLINE-5-CARBOXYLATE SYNTHETASE 1 & 2 (P5CS1 & 2) in Arabidopsis anthers and both isoforms were strongly expressed in developing microspores and pollen grains but only inconsistently in surrounding sporophytic tissues. We introduced in a p5cs1/p5cs1 p5cs2/P5CS2 mutant background an additional copy of P5CS2 under the control of the Cauliflower Mosaic Virus (CaMV) 35S promoter, the tapetum-specific LIPID TRANSFER PROTEIN 12 (Ltp12) promoter or the pollen-specific At5g17340 promoter to determine in which site proline biosynthesis can restore the fertility of proline-deficient microspores. The specificity of these promoters was confirmed by β-glucuronidase (GUS) analysis, and by direct proline measurement in pollen grains and stage-9/10 anthers. Expression of P5CS2 under control of the At5g17340 promoter fully rescued proline content and normal morphology and fertility of mutant pollen. In contrast, expression of P5CS2 driven by either the Ltp12 or CaMV35S promoter caused only partial restoration of pollen development with little effect on pollen fertility.

Conclusions

Overall, our results indicate that proline transport is not able to fulfill the demand of the cells of the male germ line. Pollen development and fertility depend on local proline biosynthesis during late stages of microspore development and in mature pollen grains.

     

Changes in plasma membrane aquaporin gene expression under osmotic stress and blue light in tomato

Divergent abiotic stresses induce osmotic stress on plant cells resulting in an imbalance in water homeostasis which is preserved by aquaporins. Since the plasma membrane aquaporins (PIPs) were shown to be involved in seed development and responses to abiotic stresses, we focused on determining the contribution of mannitol-induced osmotic stress, blue light (BL), and 7B-1 mutation to their gene expression in tomato (Solanum lycopersicum L.) seeds. To assess that, we used a quantitative RT-PCR to determine the expression profiles of genes encoding PIPs. Subsequently, a multiple linear regression analysis was used to evaluate the impact of studied stressors (mannitol and BL) and 7B-1 mutation on PIP gene expressions. We found that mannitol-induced osmotic stress and 7B-1 mutation (conferring the lower responsiveness to osmotic stress- and BL-induced inhibition of seed germination) decreased expression of PIP1;3, PIP2;3 and PIP1;2, PIP2;1 genes, respectively. This might be a way to retain water for radicle elongation and seed germination under the stress conditions. Interestingly, the expression of PIP1;3 gene was downregulated not only by osmotic stress, but also by BL. Altogether, our data indicate the existence of a link between osmotic stress and BL signalling and the involvement of the 7B-1 mutation in this crosstalk.     

Genetic control of fasciation in pea (<Emphasis Type="Italic">Pisum sativum</Emphasis> L.)

The inheritance and manifestation of fasciation character in three fasciated lines of common pea Pisum sativum L. were investigated. All studied forms are characterized by abnormal enlargement of stem apical meristem leading to distortions in shoot structure. It was estimated that fasciation in mutant Shtambovyi is connected with recessive mutation in gene FAS, which was localized in linkage group III using morphological and molecular markers. It was demonstrated that fasciation in cultivar Rosacrone and line Lupinoid is caused by recessive mutation of the same gene (FA). The peculiar architecture of inflorescence in the Lupinoid line is a result of interaction of two recessive mutations (det fa). Investigation of interaction of mutations fa and fas revealed that genes FA and FAS control consequential stages of apical meristem specialization. Data on incomplete penetrance and varying expressivity were confirmed for the mutant allele fa studied.     

Effects of interaction of <Emphasis Type="Italic">alc</Emphasis> (alcobaca) keeping-life gene with elevated fruit pigmentation genes

Results of research on the study of the effects of the interaction between the keeping-life gene alc with the elevated fruit pigmentation genes hp, dg, B ^og, and B ^c are presented. It is shown that use of the gene recombinations alc/alc//hp/hp//B ^og/B ^og(B ^c/B ^c) and alc/alc//dg/dg is the most effective means of creating highly commercial, long keeping life varieties of tomato with saturated-red coloring of the fruit.     

Genetic analysis of the traits introgressed from <Emphasis Type="Italic">Aegilops speltoides</Emphasis> Tausch. to bread wheat and determined by chromosome 5A genes

A winter bread wheat accession from the Arsenal collection was genetically examined to study the results of introgression, which substantially changed the physiological and morphological traits of the original spring cultivar Rodina. Apart from its winter habit, the accession was characterized by awned speltoid spikes, suggesting introgression into chromosome 5A, which carries marker genes in the order Vrn-A1-Q-B1. Genetic analysis showed that the chromosome fragment introgressed from Aegilops speltoides recombined well with the homeologous region of bread wheat chromosome 5A in the region between the Vrn-A1 and Q genes. Recombination between the Vrn-A1 and B1 genes was not detected, and it was assumed that the order of the marker genes of chromosome 5A was inverted to produce Q-Vrn-A1-B1. When the winter introgression line was crossed with Triticum spelta L., an interaction of two dominant genes determining the spike character was for the first time detected in F₁, increasing the spike length and the number of spikelets, and followed with transgression in F₂. It was assumed that Ae. speltoides had a homeoallelic speltoid gene, which was designated as Q ^S .