Species divergence and phylogeography of Corylus heterophylla Fisch complex (Betulaceae): Inferred from molecular, climatic and morphological data.

Historical geo-climatic changes have shaped the geographical distributions and genetic diversity of numerous plant taxa in East Asia, which promote species divergence and ultimately speciation. Here, we integrated multiple approaches, including molecular phylogeography, ecological niche modeling, and morphological traits to examine the nucleotide diversity and interspecific divergence within Corylus heterophylla complex (C. heterophylla, C. kweichowensis, and C. yunnanensis). These three sibling taxa harbored similar high levels of nucleotide diversity at the species level. The molecular data (SCNG and cpDNA) unanimously supported the division of C. heterophylla complex into two major clades, with C. yunnanensis diverged earlier from the complex, whereas C. heterophylla and C. kweichowensis could hardly be separated. The split between the two clades (c. 12.89 Ma) coincided with the formation of Sichuan Basin in the middle Miocene, while the divergence among and within the five subclades (YUN1-YUN3, HK1-HK2) occurred from the late Miocene to the Pleistocene. C. heterophylla of northern China experienced glacial contraction and interglacial expansion during the Quaternary, whereas C. kweichowensis and C. yunnanensis of southern China presented population expansion even during the last glacial maximum. Despite of high levels of genetic admixture between C. heterophylla and C. kweichowensis, significant ecological and morphological discrepancy as well as incomplete geographic isolation indicated that adaptive evolution triggered by divergent selection may have played important roles in incipient ecological speciation.

Resolving the Speciation Patterns and Evolutionary History of the Intercontinental Disjunct Genus Corylus (Betulaceae) Using Genome-Wide SNPs.

Understanding the underlying mechanisms of species origin, divergence, and distribution patterns of the intercontinental disjunct taxa has long fascinated botanists. Based on 4,894 genome-wide single-nucleotide polymorphism dataset, we present a molecular phylogenetic reconstruction of genus Corylus (Betulaceae), which have a disjunct distribution between Eurasia and North America (NA). The aim is to explore the speciation patterns and evolutionary relationships of Corylus species by establishing a general phylogenetic framework with extensive sampling. Both the molecular phylogeny inferred from recombination-free dataset and structure analysis support the division of Corylus into four major clades (A-D). Recombination tests and hybridization detection reveal extensive recombination and hybridization events among different clades, which have potentially influenced the speciation process of Corylus. Divergence time estimation indicates that recent common ancestor (MRCA) of Corylus occurred in late Eocene (∼36.38 Ma) and subsequent rapid diversification began during Miocene. Ancestral area reconstruction shows that Corylus originated from southwest China. The arrival of two clades (Clades B and C) to NA was well supported by the long distance dispersal crossing the Bering land bridge. The Himalayas, European-Mediterranean area, and other distribution regions are primarily the recipients of dispersal taxa. Vicariance after dispersal plays an important role in speciation.

Assessment of Genetic Diversity and Population Genetic Structure of Corylus mandshurica in China Using SSR Markers.

Corylus mandshurica, also known as pilose hazelnut, is an economically and ecologically important species in China. In this study, ten polymorphic simple sequence repeat (SSR) markers were applied to evaluate the genetic diversity and population structure of 348 C. mandshurica individuals among 12 populations in China. The SSR markers expressed a relatively high level of genetic diversity (Na = 15.3, Ne = 5.6604, I = 1.8853, Ho = 0.6668, and He = 0.7777). According to the coefficient of genetic differentiation (Fst = 0.1215), genetic variation within the populations (87.85%) were remarkably higher than among populations (12.15%). The average gene flow (Nm = 1.8080) significantly impacts the genetic structure of C. mandshurica populations. The relatively high gene flow (Nm = 1.8080) among wild C. mandshurica may be caused by wind-pollinated flowers, highly nutritious seeds and self-incompatible mating system. The UPGMA (unweighted pair group method of arithmetic averages) dendrogram was divided into two main clusters. Moreover, the results of STRUCTURE analysis suggested that C. mandshurica populations fell into two main clusters. Comparison of the UPGMA dendrogram and the Bayesian STRUCTURE analysis showed general agreement between the population subdivisions and the genetic relationships among populations of C. mandshurica. Group I accessions were located in Northeast China, while Group II accessions were in North China. It is worth noting that a number of genetically similar populations were located in the same geographic region. The results further showed that there was obvious genetic differentiation among populations from Northeast China to North China. Results from the Mantel test showed a weak but still significant positive correlation between Nei's genetic distance and geographic distance (km) among populations (r = 0.419, P = 0.005), suggesting that genetic differentiation in the 12 C. mandshurica populations might be related to geographic distance. These data provide comprehensive information for the development of conservation strategies of these valuable hazelnut resources.

Expression and Functional Analysis of WRKY Transcription Factors in Chinese Wild Hazel, Corylus heterophylla Fisch.

Plant WRKY transcription factors are known to regulate various biotic and abiotic stress responses. In this study we identified a total of 30 putative WRKY unigenes in a transcriptome dataset of the Chinese wild Hazel, Corylus heterophylla, a species that is noted for its cold tolerance. Thirteen full-length of these ChWRKY genes were cloned and found to encode complete protein sequences, and they were divided into three groups, based on the number of WRKY domains and the pattern of zinc finger structures. Representatives of each of the groups, Unigene25835 (group I), Unigene37641 (group II) and Unigene20441 (group III), were transiently expressed as fusion proteins with yellow fluorescent fusion protein in Nicotiana benthamiana, where they were observed to accumulate in the nucleus, in accordance with their predicted roles as transcriptional activators. An analysis of the expression patterns of all 30 WRKY genes revealed differences in transcript abundance profiles following exposure to cold, drought and high salinity conditions. Among the stress-inducible genes, 23 were up-regulated by all three abiotic stresses and the WRKY genes collectively exhibited four different patterns of expression in flower buds during the overwintering period from November to April. The organ/tissue related expression analysis showed that 18 WRKY genes were highly expressed in stem but only 2 (Unigene9262 and Unigene43101) were greatest in male anthotaxies. The expression of Unigene37641, a member of the group II WRKY genes, was substantially up-regulated by cold, drought and salinity treatments, and its overexpression in Arabidopsis thaliana resulted in better seedling growth, compared with wild type plants, under cold treatment conditions. The transgenic lines also had exhibited higher soluble protein content, superoxide dismutase and peroxidase activiety and lower levels of malondialdehyde, which collectively suggets that Unigene37641 expression promotes cold tolerance.

[Effects of saline-alkali stress on electrical impedance spectroscopy parameters and ion contents in shoots of Ping'ou hybrid hazelnut].

To study the adaptability to salt-alkaline stress of Ping'ou hybrid hazelnut, 'Liaozhen 3' shoots which were treated with three types of stress neutral NaCl, alkaline Na2CO3, and mixed salt-alkali, and the changes in electrical impedance spectroscopy (EIS) parameters and mineral ion contents were subsequently determined. The correlations between the EIS parameters and mineral ion contents were analyzed. The results showed that with the increasing level of NaCl, specific high- frequency resistance (r), specific low-frequency resistance ( r(l)), specific intracellular resistance (r(i)) and specific extracellular resistance (r(e)) of shoots decreased firstly, then increased, and finally decreased again. However, these parameters increased gradually with the increasing level of Na2CO3, while r(l) and r(e) decreased slowly in the mixed salt-alkali treatments. The Na+ contents of shoots increased significantly under the three salt-alkali stresses with the order of NaCl stress > mixed salt-alkali stress > Na2CO3 stress. Furthermore, Na2CO3 stress resulted in the decreases in the contents of three elements Zn, B and Ca. The significant negative correlation was found between the sum of five cations and four EIS parameters r(l), r(e), relaxation time (τ) , and distribution coefficient of relaxation time (ψ). The shoots of 'Liaozhen 3' might be tolerant of Na2CO3 stress of 200 mmol · L(-1), while they could be resistant to NaCl stress of 100-150 mmol · L(-1).

Vascular anatomy of kiwi fruit and its implications for the origin of carpels.

Kiwi fruit is of great agricultural, botanical, and economic interest. The flower of kiwi fruit has axile placentation, which is typical for Actinidiaceae. Axile placentation is thought derived through fusion of conduplicate carpels with marginal placentation according to the traditional doctrine. Recent progress in angiosperm systematics has refuted this traditional doctrine and placed ANITA clade rather than Magnoliaceae as the basalmost clade. However, the former traditional doctrine stays in the classrooms as the only teachable theory for the origin of carpels. To test the validity of this doctrine, we performed anatomical study on kiwi fruit. Our study indicates that the placenta has a vascular system independent of that of the ovary wall, the ovules/seeds are attached to the placenta that is a continuation of floral axis enclosed by the lateral appendages that constitute the ovary wall, and there are some amphicribral bundles in the center of placenta and numerous amphicribral bundles supplying ovules/seeds in kiwi fruit. The amphicribral vascular bundles supplying the ovules/seeds are comparable to those usually seen in branches, but not comparable to those seen in leaves or their derivatives. This comparison indicates that the placenta in kiwi fruit cannot be derived from the fusion of collateral ventral bundles of conduplicate carpels, as suggested by traditional doctrine. Instead the vascular organization in placenta of kiwi suggests that the placenta is a shoot apex-bearing ovules/seeds laterally. This conclusion is in line with the recently raised Unifying Theory, in which the placenta is taken as an ovule-bearing branch independent of the ovary wall (carpel in strict sense). Similar vascular organization in placenta has been seen in numerous isolated taxa besides kiwi fruit. Therefore whether such a pattern is applicable for other angiosperms is an interesting question awaiting answering.

[Methylenetetrahydrofolate reductase and methionine synthase reductase gene polymorphisms in ethnic Han women from Linyi].

OBJECTIVE: To explore the distribution of genetic polymorphisms of methylenetetrahydrofolate reductase (MTHFR) 677C/T, 1298A/C and methionine synthase reductase (MTRR) 66A/G among ethnic Han females from Linyi, and to correlate it with serum level of homocysteine (Hcy). METHODS: A cross-sectional study was carried out. Oral epithelial cell samples were collected from 825 subjects. MTHFR and MTRR gene polymorphisms were determined with a Taqman-Minor Groove Binder (MGB) method. Distribution of gene polymorphisms was analyzed and compared with others regions of China including Weifang, Zhengzhou, Deyang and Hainan. A biochemical assay was also carried out to determine the total Hcy in plasma of 281 subjects. The reductase activity of MTHFR was classified into decreased and stable groups according to genetic polymorphism of MTHFR. Correlation between MTHFR groups and total Hcy level were also explored. RESULTS: (1) The frequencies of MTHFR677CC, CT and TT genotypes of the selected subjects were 16.7%, 48.3% and 35.0%, respectively. The frequencies of MTHFR 1298AA, AC and CC genotypes were 76.0%, 21.6% and 2.4%, respectively. And those of MTRR 66AA, AG and GG genotypes were 54.7%, 39.4% and 5.9%, respectively. For the selected subjects, their frequency of MTHFR 677TT genotype was higher than that of Deyang and Hainan (P< 0.01), whilst the frequency of MTHFR 1298CC genotype was lower than that of Deyang and Hainan (P < 0.01), and the frequency of MTRR 66 GG genotype was lower than that of Hainan (P< 0.01). (2) The Hcy level for those with decreased MTHFR activity was significantly higher than those with stable MTHFR activity (P< 0.05). CONCLUSION: MTHFR gene 677C/T, 1298A/C and MTRR 66A/G polymorphisms in ethnic Han women from Linyi have differed significantly from other regions of China. Decreased MTHFR activity caused by genetic polymorphisms is a risk factor for raised Hcy level.