The complete chloroplast genome sequencing analysis revealed an unusual IRs reduction in three species of subfamily Zygophylloideae.

Tetraena mongolica, Zygophyllum xanthoxylon, and Z. fabago are three typical dryland plants with important ecological values in subfamily Zygophylloideae of Zygophyllaceae. Studies on the chloroplast genomes of them are favorable for understanding the diversity and phylogeny of Zygophyllaceae. Here, we sequenced and assembled the whole chloroplast genomes of T. mongolica, Z. xanthoxylon, and Z. fabago, and performed comparative genomic and phylogenetic analysis. The total size, structure, gene content and orders of these three chloroplast genomes were similar, and the three chloroplast genomes exhibited a typical quadripartite structure with a large single-copy region (LSC; 79,696-80,291 bp), a small single-copy region (SSC; 16,462-17,162 bp), and two inverted repeats (IRs; 4,288-4,413 bp). A total of 107 unique genes were identified from the three chloroplast genomes, including 70 protein-coding genes, 33 tRNAs, and 4 rRNAs. Compared with other angiosperms, the three chloroplast genomes were significantly reduced in overall length due to an unusual 16-24 kb shrinkage of IR regions and loss of the 11 genes which encoded subunits of NADH dehydrogenase. Genome-wide comparisons revealed similarities and variations between the three species and others. Phylogenetic analysis based on the three chloroplast genomes supported the opinion that Zygophyllaceae belonged to Zygophyllales in Fabids, and Z. xanthoxylon and Z. fabago belonged to Zygophyllum. The genome-wide comparisons revealed the similarity and variations between the chloroplast genomes of the three Zygophylloideae species and other plant species. This study provides a valuable molecular biology evidence for further studies of phylogenetic status of Zygophyllaceae.

Population genetic structure and gene flow of rare and endangered Tetraena mongolica Maxim. revealed by reduced representation sequencing.

BACKGROUND: Studying population genetic structure and gene flow of plant populations and their influencing factors is of particular significance in the field of conservation biology, especially important for species such as rare and endangered plants. Tetraena mongolica Maxim. (TM), belongs to Zygophyllaceae family, a rare and endangered plant with narrow distribution. However, for the last decade, due to excessive logging, urban expansion, industrial and tourism development, habitat fragmentation and loss of natural habitats have become major threats to the population of endangered plants. RESULTS: In this study, genetic diversity, population genetic structure and gene flow of TM populations were evaluated by reduced representation sequencing technology, and a total of more than 133.45 GB high-quality clean reads and 38,097 high-quality SNPs were generated. Analysis based on multiple methods, we found that the existing TM populations have moderate levels of genetic diversity, and very low genetic differentiation as well as high levels of gene flow between populations. Population structure and principal coordinates analysis showed that 8 TM populations can be divided into two groups. The Mantel test detected no significant correlation between geographical distances and genetic distance for the whole sampling. Moreover, the migration model indicated that the gene flow is more of a north to south migration pattern in history. CONCLUSIONS: This study demonstrates that the present genetic structure is mainly due to habitat fragmentation caused by urban sprawl, industrial development and coal mining. Our recommendation with respect to conservation management is that, all 8 populations should be preserved as a whole population, rather than just those in the core area of TM nature reserve. In particular, the populations near the edge of TM distribution in cities and industrial areas deserve our special protection.

Identification of Genic SSRs Provide a Perspective for Studying Environmental Adaptation in the Endemic Shrub Tetraena mongolica.

Tetraena mongolica is a xerophytic shrub endemic to desert regions in Inner Mongolia. This species has evolved distinct survival strategies that allow it to adapt to hyper-drought and heterogeneous habitats. Simple sequence repeats (SSRs) may provide a molecular basis in plants for fast adaptation to environmental change. Thus, identifying SSRs and their possible effects on gene behavior has the potential to provide valuable information for studies of adaptation. In this study, we sequenced six individual transcriptomes of T. mongolica from heterogeneous habitats, focused on SSRs located in genes, and identified 811 polymorphic SSRs. Of the identified SSRs, 172, 470, and 76 were located in 5' UTRs, CDSs, and 3' UTRs in 591 transcripts; and AG/CT, AAC/GTT, and AT/AT were the most abundant repeats in each gene region. Functional annotation showed that many of the identified polymorphic SSRs were in genes that were enriched in several GO terms and KEGG pathways, suggesting the functional significance of these genes in the environmental adaptation process. The identification of polymorphic genic SSRs in our study lays a foundation for future studies investigating the contribution of SSRs to regulation of genes in natural populations of T. mongolica and their importance for adaptive evolution of this species.

Does breeding system affect pollen morphology? A case study in Zygophylloideae (Zygophyllaceae).

KEY MESSAGE: We found a correlation between large pollens and selfing and between psilate pollen walls and cleistogamy. Flowering plants display extraordinary diversity in pollen morphology, yet the functional significance of this variation is not well understood. Zygophylloideae is a lineage characterized by high diversity of breeding systems, ranging from obligate selfing to facultative selfing and further to facultative outcrossing. This group is particularly suitable for testing hypotheses about the influence of breeding system on pollen morphology. We studied pollen morphology in 20 species of Zygophylloideae and one species of Tribuloideae as an outgroup. A phylogeny of Zygophylloideae was created based on available DNA sequences and used to trace the evolution of pollen characters. We performed a phylogenetic analysis of correlated evolution between breeding system and several pollen characters. Three types of pollen morphology were found in the studied species. Tricolporate pollen with a small apocolpium, microreticulate ornamentation and medium size was determined as the ancestral state in Zygophylloideae. The correlation analysis indicated an association between large pollens and selfing and between psilate pollen wall and cleistogamy. We hypothesize that large size of pollen in selfing species is mainly associated with the low number of produced pollen indicating a trade-off between pollen size and number. The independence from pollen vector in cleistogamous flowers accounts for the evolution of smooth pollen walls in these species.

The assessment of cadmium, chromium, copper, and nickel tolerance and bioaccumulation by shrub plant Tetraena qataranse.

Heavy metals constitute some of the most significant environmental contaminants today. The abundance of naturally growing Tetraena qataranse around Ras Laffan oil and gas facilities in the state of Qatar reflects its toxitolerant character. This study examined the desert plant's tolerance to Ba, Cd, Cr, Cu, Ni and Pb relative to soil concentration. Analysis by inductively coupled plasma - optical emission spectroscopy (ICP-OES) showed that the plant biomass accumulates higher Cd, Cr, Cu and Ni concentration than the soil, particularly in the root. The bioconcentration factor (BCF) of all metals in the root and shoot indicates the plant's capacity to accumulate these metals. Cd had a translocation factor (TF) greater than one; however, it is less than one for all other metals, suggesting that the plant remediate Cd by phytoextraction, where it accumulates in the shoot and Cr, Cu and Ni through phytostabilization, concentrating the metals in the root. Metals phytostabilization restrict transport, shield animals from toxic species ingestion, and consequently prevent transmission across the food chain. Fourier Transform Infrared Spectroscopy (FTIR) analysis further corroborates ICP-OES quantitative data. Our results suggest that T. qataranse is tolerant of Cd, Cr, Cu, and Ni. Potentially, these metals can accumulate at higher concentration than shown here; hence, T. qataranse is a suitable candidate for toxic metals phytostabilization.

Evolution of leaf anatomy in arid environments - A case study in southern African Tetraena and Roepera (Zygophyllaceae).

The dry biomes of southern Africa (Desert, Nama Karoo and Succulent Karoo) are home to a rich and diverse xerophytic flora. This flora includes two morphologically diverse clades of Zygophyllaceae, Tetraena and Roepera (Zygophylloideae), which inhabit some of the most arid habitats in the region. Using a plastid phylogeny of Zygophylloideae we assess whether the evolution of putatively adaptive traits (leaf shape, vasculature, mode of water storage and photosynthetic type: C3 versus C4) coincides with the successful colonisation of environments with different drought regimes within southern Africa. Our results show general niche conservatism within arid habitats in Tetraena, but niche shifts from arid to more mesic biomes with longer and/or cooler growing season (Fynbos and Thicket) in Roepera. However, these distinct broad-scale biogeographical patterns are not reflected in leaf anatomy, which seems to vary at more local scales. We observed considerable variability and multiple convergences to similar leaf anatomies in both genera, including shifts between "all cell succulence" leaf types and leaf types with distinct chlorenchyma and hydrenchyma. Our survey of C4 photosynthesis in the Zygophylloideae showed that the C4 pathway is restricted to Tetraena simplex, which also having an annual life history and a widespread distribution, is rather atypical for this group.

Evolution of Asian Interior Arid-Zone Biota: Evidence from the Diversification of Asian Zygophyllum (Zygophyllaceae).

The Asian interior arid zone is the largest desert landform system in the Northern Hemisphere, and has high biodiversity. Little is currently known about the evolutionary history of its biota. In this study, we used Zygophyllum, an important and characteristic component of the Asian interior arid zone, to provide new insights into the evolution of this biota. By greatly enlarged taxon sampling, we present the phylogenetic analysis of Asian Zygophyllum based on two plastid and one nuclear markers. Our phylogenetic analyses indicate that Asian Zygophyllum and Sarcozygium form a clade and Sarcozygium is further embedded within the shrub subclade. An integration of phylogenetic, biogeographic, and molecular dating methods indicates that Zygophyllum successfully colonized the Asian interior from Africa in the early Oligocene, and Asian Zygophyllum became differentiated in the early Miocene and underwent a burst of diversification in the late Miocene associated with the expansion of Asian interior arid lands due to orogenetic and climatic changes. Combining diversification patterns of other important components of the Asian interior arid zone, we propose a multi-stage evolution model for this biota: the late Eocene-early Oligocene origin, the early Miocene expansion, and the middle-late Miocene rapid expansion to the whole Asian interior arid zone. This study also demonstrates that, for Zygophyllum and perhaps other arid-adapted organisms, arid biomes are evolutionary cradles of diversity.

Cloning, characterization and functional analysis of two type 1 diacylglycerol acyltransferases (DGAT1s) from Tetraena mongolica.

Two cDNAs encoding putative type 1 acyl-CoA: diacylglycerol acyltransferases (DGAT1, EC 2.3.1.20), were cloned from Tetraena mongolica, an extreme xerophyte with high oil content in the stems. The 1 488-bp and 1 485-bp of the open reading frame (ORF) of the two cDNAs, designated as TmDGAT1a and TmDGAT1b, were both predicted to encode proteins of 495 and 494 amino acids, respectively. Southern blot analysis revealed that TmDGAT1a and TmDGAT1b both had low copy numbers in the T. mongolica genome. In addition to ubiquitous expression with different intensity in different tissues, including stems, leaves and roots, TmDGAT1a and TmDGAT1b, were found to be strongly induced by high salinity, drought and osmotic stress, resulting in a remarkable increase of triacylglycerol (TAG) accumulation in T. mongolica plantlets. TmDGAT1a and TmDGAT1b activities were confirmed in the yeast H1246 quadruple mutant (DGA1, LRO1, ARE1, ARE2) by restoring DGAT activity of the mutant host to produce TAG. Overexpression of TmDGAT1a and TmDGAT1b in soybean hairy roots as well as in T. mongolica calli both resulted in an increase in oil content (ranging from 37% to 108%), accompanied by altered fatty acid profiles.

Salinity-induced changes in protein expression in the halophytic plant Nitraria sphaerocarpa.

Salinity is a major abiotic stress that inhibits plant growth and development. Plants have evolved complex adaptive mechanisms that respond to salinity stress. However, an understanding of how plants respond to salinity stress is far from being complete. In particular, how plants survive salinity stress via alterations to their intercellular metabolic networks and defense systems is largely unknown. To delineate the responses of Nitraria sphaerocarpa cell suspensions to salinity, changes in their protein expression patterns were characterized by a comparative proteomic approach. Cells that had been treated with 150 mM NaCl for 1, 3, 5, 7, or 9 days developed several stress-related phenotypes, including those affecting morphology and biochemical activities. Of ~1100 proteins detected in 2-DE gel patterns, 130 proteins showed differences in abundance with more than 1.5-fold when cells were stressed by salinity. All but one of these proteins was identified by MS and database searching. The 129 spots contained 111 different proteins, including those involved in signal transduction, cell rescue/defense, cytoskeleton and cell cycle, protein folding and assembly, which were the most significantly affected. Taken together, our results provide a foundation to understand the mechanism of salinity response.

Conservation genetics and phylogeography of endangered and endemic shrub Tetraena mongolica (Zygophyllaceae) in Inner Mongolia, China.

BACKGROUND: Tetraena mongolica (Zygophyllaceae), an endangered endemic species in western Inner Mongolia, China. For endemic species with a limited geographical range and declining populations, historical patterns of demography and hierarchical genetic structure are important for determining population structure, and also provide information for developing effective and sustainable management plans. In this study, we assess genetic variation, population structure, and phylogeography of T. mongolica from eight populations. Furthermore, we evaluate the conservation and management units to provide the information for conservation. RESULTS: Sequence variation and spatial apportionment of the atpB-rbcL noncoding spacer region of the chloroplast DNA were used to reconstruct the phylogeography of T. mongolica. A total of 880 bp was sequenced from eight extant populations throughout the whole range of its distribution. At the cpDNA locus, high levels of genetic differentiation among populations and low levels of genetic variation within populations were detected, indicating that most seed dispersal was restricted within populations. CONCLUSIONS: Demographic fluctuations, which led to random losses of genetic polymorphisms from populations, due to frequent flooding of the Yellow River and human disturbance were indicated by the analysis of BEAST skyline plot. Nested clade analysis revealed that restricted gene flow with isolation by distance plus occasional long distance dispersal is the main evolutionary factor affecting the phylogeography and population structure of T. mongolica. For setting a conservation management plan, each population of T. mongolica should be recognized as a conservation unit.

Genetic diversity among Zygophyllum (Zygophyllaceae) populations based on RAPD analysis.

Zygophyllum species are succulent plants that are drought resistant and/or salt tolerant, growing under severe, dry climatic conditions. Despite their importance and abundance in the Mediterranean and Middle East regions, there is little information concerning molecular variations among species of this genus. Genetic diversity was assessed, using RAPD primers, of 12 populations of Z. coccineum, Z. album and Z. aegyptium collected from various locations in Egypt and Saudi Arabia. Yong leaves were used for DNA extraction. Genetic distances were calculated using Nei's method. A dendrogram was constructed based on the similarity data matrix by unweighted pair group method using arithmetic averages cluster analysis. Analysis with RAPD markers revealed genetic variation between and within populations of Zygophyllum. Zygophyllum coccineum showed higher levels of genetic variation and more unique alleles than the other species.

Phylogenetic relationships, character evolution and biogeography of southern African members of Zygophyllum (Zygophyllaceae) based on three plastid regions.

The plastid coding rbcL and non-coding trnLF regions of 53 of 55 southern African Zygophyllum species were sequenced and used to evaluate the phylogenetic relationships within the southern African representatives of the genus. Published sequences of the same gene regions of Australian, Asian and North African Zygophyllum species were included to assess the relationships of the species from these regions to the southern African species. The addition of Z. stapffii from Namibia, found to be conspecific with Z. orbiculatum from Angola, lead to a greatly resolved tree. The molecular results were largely congruent with a recent sectional classification of the southern African species and supported their subdivision into subgenera Agrophyllum and Zygophyllum. Reconstruction of the character evolution of capsule dehiscence, seed attachment and seed mucilage showed that these characters allowed a division of southern African species into the two subgenera but that this could not be applied to species occurring elsewhere. Other morphological characters were found to vary and unique character combinations, rather than unique characters, were found to be of systematic value in sectional delimitation. The study suggests that repeated radiations from the horn of Africa to southern Africa and Asia and back lead to the present distribution of the taxa in the subfamily Zygophylloideae. Although this study supports some of the recent taxonomic changes in the group, the unresolved relationships between the proposed genera Tetraena and Roepera and those retained as Zygophyllum species suggest that changes to the taxonomy may have been premature.

The genetic structure of the gynodioecious Kallstroemia grandiflora (Zygophyllaceae): the role of male sterility and colonization history.

In gynodioecious populations, the frequency of females is expected to have a strong influence on the contemporary genetic structure of populations. Historical patterns of range contraction and expansion are also known to influence the genetic diversity of plant populations. We explore the influence of male sterility and colonization history on the genetic diversity in populations of Kallstroemia grandiflora along the Pacific of México. Both the overall population Fis and Fis values of hermaphrodites showed a negative association with female frequency. Genetic diversity declined with latitude. Our results provide evidence that females have a significant effect on the genetic structure as predicted by theoretical models and provide support for the hypothesis that historical processes have modified the genetic structure of K. grandiflora.

Phylogenetic relationships and biogeography of the desert plant genus Fagonia (Zygophyllaceae), inferred by parsimony and Bayesian model averaging.

Phylogenetic relationships within Fagonia were inferred from analyses of plastid trnL intron and nuclear ribosomal ITS DNA sequences. Sampling of the genus was nearly complete, including 32 of 34 species. Phylogenetic analysis was carried out using parsimony, and Bayesian model averaging. The latter method allows model-based inference while accounting for model-selection uncertainty, and is here used for the first time in phylogenetic analyses. All species of Fagonia in the Old World, except F. cretica, form a weakly supported clade, and all Fagonia species of the New World, except F. scoparia, are well supported as sister to the Old World clade. Fagonia scoparia, from Mexico, and F. cretica, from Northern Africa, are well supported as sisters to all other Fagonia species. Vicariance-dispersal analysis, using DIVA, indicated that the occurrences of Fagonia in South America and southern Africa are due to dispersals, and also, that the ancestor of Fagonia had a distribution compatible with the boreotropics hypothesis.