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Mountain systems harbor an evolutionarily unique and exceptionally rich biodiversity, especially for amphibians. However, the associated elevational gradients and underlying mechanisms of amphibian diversity in most mountain systems remain poorly understood. Here, we explored amphibian phylogenetic and functional diversity along a 2 600 m elevational gradient on Mount Emei on the eastern margin of the Qinghai-Tibetan Plateau in southwestern China. We also assessed the relative importance of spatial (area) and environmental factors (temperature, precipitation, solar radiation, normalized difference vegetation index, and potential evapotranspiration) in shaping amphibian distribution and community structure. Results showed that the phylogenetic and functional diversities were unimodal with elevation, while the standardized effect size of phylogenetic and functional diversity increased linearly with elevation. Phylogenetic net relatedness, nearest taxon index, and functional net relatedness index all showed a positive to negative trend with elevation, indicating a shift from clustering to overdispersion and suggesting a potential change in key processes from environmental filtering to competitive exclusion. Overall, our results illustrate the importance of deterministic processes in structuring amphibian communities in subtropical mountains, with the dominant role potentially switching with elevation. This study provides insights into the underlying assembly mechanisms of mountain amphibians, integrating multidimensional diversity.山地系统孕育了进化上独特且极为丰富的生物多样性，对两栖动物来说尤其如此。然而，当前大多数山地系统中两栖动物沿海拔梯度的多样性格局及群落构建机制尚不清楚。峨眉山位于中国西南地区的青藏高原东缘，其丰富的生物多样性及较大的海拔跨度（2 600 m）为理解两栖动物的海拔梯度格局提供了理想的研究平台。该研究基于峨眉山两栖动物的详细调查数据，解析了其系统发育和功能多样性沿海拔梯度的分布格局及群落结构，并进一步评估了空间（面积）和环境因子（温度、降水、太阳辐射、归一化植被指数和潜在蒸发量）在不同维度两栖动物多样性及群落构建中的重要性。结果表明，峨眉山两栖动物系统发育和功能多样性沿海拔均呈单峰分布；控制物种数的影响后，它们的标准化效应值随海拔升高单调递增。系统发育净亲缘指数、最近亲缘指数及功能净亲缘指数均沿海拔呈现出由正转负的趋势，这表明群落结构经历了由聚集到发散的转变，并暗示构建机制可能存在由环境过滤到竞争排斥的变化。该研究阐明了环境过滤和竞争排斥在亚热带山地系统两栖动物群落构建中的重要性：随着海拔升高，两者的主导作用可能会发生转换。通过整合不同维度的生物多样性，该研究有助于深入理解山地系统两栖动物的群落构建机制。.

Background Alzheimer’s disease (AD) and glioblastoma are the most common and devastating diseases in the neurology and neurosurgery departments, respectively. Our previous research reports that the AD-related protein Presenilin1 represses cell proliferation by inhibiting the Wnt/β-catenin pathway in glioblastoma. However, the function of Presenilin1 and the underlying mechanism need to be further investigated. Methods The correlations of two genes were conducted on the R2 microarray platform and CGGA. Wound healing, Transwell assays and glioblastoma transplantation were performed to detect invasion ability. Phalloidin staining was employed to show cell morphology. Proximity ligation assays and protein docking assays were employed to detect two protein locations. We also employed western blotting to detect protein expression. Results We found that Presenilin1 clearly repressed the migration, invasion and mesenchymal transition of glioblastoma cells. Intriguingly, we observed that the expression of Presenilin1 was positively correlated with Sortilin, which is identified as a pro-invasion molecule in glioma. Furthermore, Presenilin1 interacted with Sortilin at the transmembrane domain and repressed Sortilin expression by cleaving it in glioblastoma cells. First, we found that Sortilin introduced the function of Presenilin1 in phosphorylating β-catenin and repressing invasion in glioblastoma cells. Last, Presenilin1 stimulation sharply suppressed the invasion and mesenchymal transition of glioblastoma in mouse subcutaneous and intracranial transplantation models. Conclusions Our study reveals that Sortilin mediates the regulation of β-catenin by Presenilin1 and transduces the anti-invasive function of Presenilin1, which may provide novel therapeutic targets for glioblastoma treatment.

Adhesion molecules play an important role in urinary calculus formation. The expressions of adhesion molecules in renal tubular has been reported in some animals. However, the role of adhesion molecules in the process of sheep urinary calculus formation is still unclear. The magnesium ammonium phosphate (MAP) is the main component of sheep urinary calculus. In this paper, the sheep renal tubular epithelial cells (RTECs) were isolated and treated with MAP, the expressions of osteopontin (OPN), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and apoptosis-related indicators caspase-3, Bcl-2 and Bax in RTECs were observed, the viability of RTECs was detected by Cell Counting Kit-8 (CCK-8). The levels of superoxide dismutase (SOD) and malondialdehyde (MDA), and the expressions of inflammatory factors Interleukin-6 (IL-6), Interleukin-1 (IL-1), Interleukin-17 (IL-17) and tumor necrosis factor-α (TNF-α) were measured by enzyme-linked immunosorbent (ELISA). The histopathological observation of kidney in urolithiasis sheep was made. The results showed that MAP could reduce the viability and SOD activity, enhance the activity of MDA significantly and promote the expressions of IL-1, IL-6, IL-17 and TNF-α of RTECs. By western blot and qPCR methods, the expressions of ICAM-1, VCAM-1 and OPN increased in 48 h. In addition, the expression of caspase-3 increased significantly and the ratio of Bcl-2/Bax reduced with exposure to MAP. The renal tissue structure was seriously damaged, the RTECs in urolithiasis sheep were degenerative and necrotic.

Embryogenic callus induction and regeneration are useful in many aspects of plant biotechnology, especially in the functional characterization of economically important genes. However, in sugarcane, callus induction and regeneration vary across genotypes.Saccharum spontaneumis an important wild germplasm that confers disease resistance and stress tolerance to modern sugarcane cultivars, and its genome has been completely sequenced. The aim of this study was to investigate the effect of genetic variations on embryogenic callus induction and regeneration inS. spontaneumand to screen genotypes having high tissue culture susceptibility. The study was performed using nine genotypes ofS. spontaneumand the following five parameters were assessed to determine the response of genotypes to embryogenic callus induction and regeneration: callus induction, embryogenic callus ratio, embryogenic callus induction, embryonic callus regeneration and regeneration capacity. All the genotypes varied significantly (P< 0.01) in all the parameters, except for embryonic callus regeneration, which was high (>80%) for all the genotypes. High broad-sense heritability (86.1–96.8%) indicated that genetic differences are the major source of genotypic variations. Callus induction was found to be strongly positively correlated with embryogenic callus induction (r= 0.890,P< 0.01) and regeneration capacity (r= 0.881,P< 0.01). Among the nine tested genotypes, VN2 was found to be the most responsive to tissue culture and could therefore be used to characterize functional genes inS. spontaneum. We also suggested an approach with potential applications in facilitating the rapid identification of sugarcane genotypes susceptible to tissue culture.

Sugarcane brown rust is a fungal disease with global distribution that is caused by Puccinia melanocephala, and is responsible for severe yield and sucrose losses in sugarcane production. The most effective strategy of controlling the disease is breeding and cultivating resistant varieties. Bru1 is a major gene that has been demonstrated to confer resistance to P. melanocephala strains from different parts of the globe. R12H16 and 9O20-F4 are two molecular markers that have been previously reported to be closely associated with Bru1, and are often used in brown rust resistance identification during sugarcane germplasm evaluation activities. In the present study, a novel type of molecular marker based on four primers is used to detect Bru1 among 200 sugarcane ancestral species and landraces from the China National Germplasm Repository of Sugarcane (NGRS). According to the results, among the 200 tested materials, 125 materials, such as 51NG63, Katha, and Uba, Bru1 could be detected based on bands corresponding to those in POJ2878, indicated that the 125 (62.5%) materials contained Bru1, and the detection rates of Bru1 in 69 sugarcane ancestral species and 131 landraces were 73.9% and 56.5%, respectively. These results indicated that the sugarcane ancestral species and landraces conserved in the China NGRS contain abundant brown rust-resistant materials, which could provide resources that could facilitate the breeding of brown rust-resistant sugarcane varieties.

The expensive tissue hypothesis predicts a trade-off between investments in the brain and other energetically costly organs due to the costs associated with their growth and maintenance within the finite energy resources available. However, few studies address the strength of relationships between brain size and investments in precopulatory (ornaments and armaments) and postcopulatory (testes and ejaculates) sexual traits. Here, in a broad comparative study, we tested the prediction that the relationship between brain size and investment in sexual traits differs among taxa relative to the importance of sperm competition within them. We found that brain size was negatively correlated with sexual size dimorphism (SSD) in anurans and primates, and it tended to decrease with SSD in ungulates and cetaceans. However, brain size did not covary significantly with armaments (e.g., canine length, horn, antler, and muscle mass). Brain size was not correlated with postcopulatory sexual traits (testes and ejaculates). The intensity of covariance between brain size and precopulatory sexual traits decreased with increasing relative testis size.

The invasion ability of glioblastoma (GBM) causes tumor cells to infiltrate the surrounding brain parenchyma and leads to poor outcomes. Transient receptor potential vanilloid 4 (TRPV4) exhibits a remarkable role in cancer cell motility, but the contribution of TRPV4 to glioblastoma metastasis is not fully understood. Here, we reported that TRPV4 expression was significantly elevated in malignant glioma compared to normal brain and low-grade glioma, and TRPV4 expression was negatively correlated with the prognosis of glioma patients. Functionally, stimulation of TRPV4 promoted glioblastoma cell migration and invasion, and repression of TRPV4 hindered the migration and invasion of glioblastoma cells in vitro. Molecularly, TRPV4 strongly colocalized and interacted with skeletal protein-F-actin at cellular protrusions, and TRPV4 regulated the formation of invadopodia and filopodia in glioblastoma cells. Furthermore, the Cdc42/N-wasp axis mediated the effect of TRPV4-regulated cellular protrusions and invasion. Foremost, TRPV4 inhibitor treatment or downregulation of TRPV4 significantly reduced the invasion-growth of subcutaneously and intracranially transplanted glioblastoma in mice. In conclusion, the TRPV4/Cdc42/wasp signaling axis regulates cellular protrusion formation in glioblastoma cells and influences the invasion-growth phenotype of glioblastoma in vivo. TRPV4 may serve as a prognostic factor and specific therapeutic target for GBM patients.

The use of nectar-providing plants to nourish natural enemies of pest species has become a widely-used approach in conservation biological control to reduce pest damage without the indiscriminate use of insecticides. Choice of plant species is crucial to maximize benefits, but suitable species are yet to be identified for many important crop-pest systems. Here we explored the suitability of three candidate nectar plants for use in brassica vegetables to suppress the globally significant pest, Plutella xylostella L. (Lepidoptera: Plutellidae), using the widely-distributed parasitoid, Cotesia vestalis (Haliday) (Hymenoptera: Braconidae). Volatiles of alyssum (Lobularia maritima (L.) Desv) (Brassicaceae) were attractive to the parasitoid and access to flowering shoots increased adult longevity and realized fecundity of C. vestalis. Moreover, adult diamondback moth derived no benefit from this flower. In contrast, buckwheat (Fagopyrum esculentum Moench) (Polygonaceae), a species widely used in conservation biological control in other systems, increased the longevity and fecundity of both pest and parasitoid, rendering it less suitable. A third plant, heronsbill (Portulaca grandiflora Hook.) (Portulacaceae) denied benefit to the pest and promoted longevity of the parasitoid under no-choice conditions but did not improve fecundity and was repellent to female parasitoids under choice conditions. The contrasting effects of this set of plants illustrate the need to test multiple response variables and effects on both pest and natural enemy when seeking optimal nectar plants for use in a novel conservation biological control system.

While crypsis is a prominent antipredator adaptation, the role of the brain in predator-driven evolution remains controversial. Resolving this controversy requires contextualizing the brain with established antipredator traits and predation pressure. We hypothesize that the reduced predation risk through crypsis relaxes predation-driven selection on the brain and provide comparative evidence across 102 Chinese frog species for our hypothesis. Specifically, our phylogenetic path analysis reveals an indirect relationship between predation risk and crypsis that is mediated by brain size. This result suggests that at a low predation risk, frogs can afford to be conspicuous and use their large brain for cognitive predator evasion. This strategy may become less efficient or energetically costlier under higher predation pressure, favoring smaller brains and instead increasing crypsis.

Altricial birds show enormous intraspecific diversity in their provisioning strategies, in terms of both the provisioning rate and the amount of food delivered per feeding bout. Extra-pair copulations (EPCs), which result in either extra-pair paternity (EPP) or maternity (EPM), provide an opportunity to demonstrate why provisioning strategies vary among individuals. Because EPP-cuckolded males and EPM-cuckolded females must raise unrelated young, whereas EPM-cuckolded males and EPP-cuckolded females need not, we hypothesized that the first two categories of breeders would reduce parental effort, whereas the latter two categories of breeders would increase parental effort. We tested this hypothesis in the Azure-winged Magpie Cyanopica cyanus by comparing the number and body mass of fledglings, provisioning rates and food amount delivered per feeding bout between EPP- and EPM-cuckolded breeders and faithful breeders. We found that (1) the number of fledglings did not differ significantly between cuckolded and faithful breeders, and (2) fledglings raised by EPM-cuckolded males and EPP-cuckolded females did not differ from faithful breeders, whereas fledglings raised by EPP-cuckolded males and EPM-cuckolded females were significantly smaller than those raised by faithful breeders. Compared with faithful breeders, cuckolded breeders increased parental efforts in nests that contained no unrelated young; hence, their loss in parentage may be compensated for by the enhanced quality of fledglings. In nests that contained unrelated young, cuckolded breeders did not reduce parental efforts; hence, their own offspring would not starve and could survive in competition with mixed brood-mates. Our findings suggest that the differences in parentage created by EPP and EPM scenarios and the potential fitness return of raising a brood for the cuckolded breeders can explain the intraspecific variation in provisioning strategies of altricial birds.

Vertebrate eye size typically scales hypoallemetrically with body size—as animals grow larger their eyes get relatively smaller. Additionally, eye size is highly variable across species, and such variability often reflects functional adaptations to differences in behavior and/or ecology. The selective pressures underlying the evolution of eye size are especially well studied in birds, mammals, and fishes. However, whether similar scaling rules and selective pressures also underlie the evolution of eye size in amphibians remains enigmatic. Variation in eye size is intimately linked with variation in brain anatomy, as the retina is ontogenetically part of the brain. Eye size may therefore coevolve with brain size. Here we use phylogenetic comparative methods to study interspecific variation in eye volume across 44 species of anurans from 8 families from the Hengduan Mountains, China. We relate this variation to key factors known to impact eye size evolution in other vertebrate taxa such as body mass, habitat use, defense strategy and foraging mobility. We found that also in anurans eyes size scaled hypoallometrically with body mass. However, neither of the behavioral or ecological factors explained any variation in relative eye size in our sample. Whether this is representative for other frog species needs to be clarified. We therefore conclude that eye size in frogs is tightly linked to body mass evolution but that, at least in the species investigated here, none of our tested ecological and behavioral factors have a strong influence on eye size evolution.

A parasitoid’s decision to reject or accept a potential host is fundamental to its fitness. Superparasitism, in which more than one egg of a given parasitoid species can deposit in a single host, is usually considered sub-optimal in systems where the host is able to support the development of only a single parasitoid. It follows that selection pressure may drive the capacity for parasitoids to recognize parasitized hosts, especially if there is a fitness cost of superparasitism. Here, we used microsatellite studies of two distinct populations of Cotesia vestalis to demonstrate that an egg laid into a diamondback moth (Plutella xylostella) larva that was parasitized by a conspecific parasitoid 10 min, 2 or 6 h previously was as likely to develop and emerge successfully as was the first-laid egg. Consistent with this, a naive parasitoid encountering its first host was equally likely to accept a healthy larva as one parasitized 10 min prior, though handling time of parasitized hosts was extended. For second and third host encounters, parasitized hosts were less readily accepted than healthy larvae. If 12 h elapsed between parasitism events, the second-laid egg was much less likely to develop. Discrimination between parasitized and healthy hosts was evident when females were allowed physical contact with hosts, and healthy hosts were rendered less acceptable by manual injection of parasitoid venom into their hemolymph. Collectively, these results show a limited capacity to discriminate parasitized from healthy larvae despite a viability cost associated with failing to avoid superparasitism.

Background Glioblastoma and Alzheimer’s disease (AD) are the most common and devastating diseases in the central nervous system. The dysfunction of Presenilin1 is the main reason for AD pathogenesis. However, the molecular function of Presenilin1 and its relative mechanism in glioblastoma remain unclear. Methods Expression of presenilin1 in glioma was determined by IHC. CCK-8, colony formation, Flow cytometry, Edu staining were utilized to evaluate functions of presenilin1 on glioblastoma proliferation. The mechanism of above process was assessed by Western blotting and cell immunofluorescence. Mouse transplanting glioblastoma model and micro-MRI detection were used to verified presenilin1 function in vivo. Results In this study, we found that all grades of glioma maintained relatively low Presenilin1 expression and that the expression of Presenilin1 in high-grade glioma was significantly lower than that in low-grade glioma. Moreover, the Presenilin1 level had a positive correlation with glioma and glioblastoma patient prognosis. Next, we determined that Presenilin1 inhibited the growth and proliferation of glioblastoma cells by downregulating CDK6, C-myc and Cyclin D1 to arrest the cell cycle at the G1/S phase. Mechanistically, Presenilin1 promoted the direct phosphorylation of β-catenin at the 45 site and indirect phosphorylation at the 33/37/41 site, then decreased the stabilized part of β-catenin and hindered its translocation from the cytoplasm to the nucleus. Furthermore, we found that Presenilin1 downregulation clearly accelerated the growth of subcutaneous glioblastoma, and Presenilin1 overexpression significantly repressed the subcutaneous and intracranial transplantation of glioblastoma by hindering β-catenin-dependent cell proliferation. Conclusion Our data implicate the antiproliferative effect of Presenilin1 in glioblastoma by suppressing Wnt/β-catenin signaling, which may provide a novel therapeutic agent for glioblastoma.

Sperm competition is often considered the primary selective force underlying the rapid and diversifying evolution of ejaculate traits. Yet, several recent studies have drawn attention to other forms of selection with the potential of exceeding the effects of sperm competition. Since ejaculates are complex, multivariate traits, it seems plausible that different ejaculate components vary in their responses to different selective pressures. Such information, however, is generally lacking as individual ejaculate traits tend to be studied in isolation. Here, we studied the macroevolutionary patterns of ejaculate volume, sperm number, sperm length and the proportion of viable normal sperm in response to varying levels of sperm competition, body size and the duration of female sperm storage in pheasants and allies (Phasianidae). Ejaculate volume, sperm number and sperm viability were all relatively higher in polygamous than in monogamous mating systems. However, whereas ejaculate volume additionally covaried with body size, sperm number instead increased with the female sperm-storage duration, in conjunction with a decrease in sperm length. Overall, our results revealed important details on how different forms of selection can jointly shape ejaculates as complex, composite traits.

Brain sizes vary substantially across vertebrate taxa, yet, the evolution of brain size appears tightly linked to the evolution of life histories. For example, larger brained species generally live longer than smaller brained species. A larger brain requires more time to grow and develop at a cost of exceeded gestation period and delayed weaning age. The cost of slower development may be compensated by better homeostasis control and increased cognitive abilities, both of which should increase survival probabilities and hence life span. To date, this relationship between life span and brain size seems well established in homoeothermic animals, especially in mammals. Whether this pattern occurs also in other clades of vertebrates remains enigmatic. Here, we undertake the first comparative test of the relationship between life span and brain size in an ectothermic vertebrate group, the anuran amphibians. After controlling for the effects of shared ancestry and body size, we find a positive correlation between brain size, age at sexual maturation, and life span across 40 species of frogs. Moreover, we also find that the ventral brain regions, including the olfactory bulbs, are larger in long-lived species. Our results indicate that the relationship between life history and brain evolution follows a general pattern across vertebrate clades.

Substantial evidence indicates that the alteration of the cellular redox status is a critical factor involved in cell growth and death and results in tumourigenesis. Cancer cells have an efficient antioxidant system to counteract the increased generation of ROS. However, whether this ability to survive high levels of ROS has an important role in the growth and metastasis of tumours is not well understood. Glutaredoxin 3 (GLRX3), also known as TXNL2, Grx3 and PICOT, maintains a low level of ROS, thus contributing to the survival and metastasis of several types of cancer. However, little is known about the role of GLRX3 and the underlying mechanisms that suppress oral squamous cell carcinoma (OSCC) progression. Here, by using immunohistochemical staining, we demonstrated that GLRX3 was overexpressed in human OSCC, and enhanced GLRX3 expression correlated with metastasis and with decreased overall patient survival. Knockdown of GLRX3 in human OSCC cell lines reduced Notch activity by reversing the epithelial-mesenchymal transition (EMT), resulting in the inhibition of in vitro migration and invasion. Importantly, knockdown of GLRX3 triggered the generation of ROS. Furthermore, N-acetyl cysteine (NAC), an ROS scavenger, enhanced the effects of GLRX3 knockdown on Notch-dependent EMT. Collectively, these findings suggested the vital roles of GLRX3 in OSCC progression through its relationship with EMT progression, and these data also suggest that a strategy of blocking ROS to enhance the activity of GLRX3 knockdown warrants further attention in the treatment of OSCC.

The ‘cognitive buffer’ hypothesis predicts that the costs of relatively large brains are compensated for later in life by the increased benefits of large brains providing a higher chance of survival under changing environments through flexible behaviors in the animal kingdom. Thus, animals that live in a larger range (with a higher probability of environmental variation) are expected to have larger brains than those that live in a restricted geographic range. Here, to test the prediction of the ‘cognitive buffer’ hypothesis that larger brains should be expected to occur in species living in geographic ranges of larger size, we analyzed the relationship between the size of the geographic range and brain size and the size of various brain regions among 42 species of anurans using phylogenetic comparative methods. The results show that there is no correlation between relative brain size and size of the species’ geographic range when correcting for phylogenetic effects and body size. Our findings suggest that the effects of the cognitive buffer and the energetic constraints on brains result in non-significant variation in overall brain size. However, the geographic range is positively correlated with cerebellum size, but not with optic tecta, suggesting that species distributed in a wider geographic range do not exhibit larger optic tecta which would provide behavioral flexibility to allow for an early escape from potential predators and discovery of new food resources in unpredictable environments.

The evolution of sperm quality and quantity is shaped by various selective processes, with sperm competition generally considered the primary selective agent. Particularly in external fertilizers, however, sperm limitation through gamete dispersal can also influence gamete investments, but empirical data examining this effect are limited. Here, we studied the relative importance of sperm competition and the spawning conditions in explaining the macroevolutionary patterns of sperm size and number within two taxa with external fertilization but differences in their reproductive biology. In frogs, sperm swim slowly but for up to hours as they penetrate the gelatinous egg coating, whereas fish sperm typically swim fast, are very short-lived (seconds to minutes), and often face a relatively higher risk of being moved away from the ova by currents. Our phylogenetic models and path analyses revealed different trajectories of ejaculate evolution in these two taxa. Sperm size and number responded primarily to variation in sperm competition in the anurans, but more strongly to egg number and water turbulence in the fishes. Whereas the results across anurans align with the general expectation that sexual selection is the main driver of ejaculate evolution, our findings across the fishes suggest that sperm limitation has been underappreciated.

The challenges of seasonal environments are thought to contribute to brain evolution, but in which way is debated. According to the Cognitive Buffer Hypothesis (CBH) brain size should increase with seasonality, as the cognitive benefits of a larger brain should help overcoming periods of food scarcity via, for instance, increased behavioral flexibility. However, in line with the Expensive Brain Framework (EBF) brain size should decrease with seasonality because a smaller brain confers energetic benefits in periods of food scarcity. Empirical evidence is inconclusive and mostly limited to homoeothermic animals. Here we used phylogenetic comparative analyses to test the impact of seasonality on brain evolution across 30 species of anurans (frogs) experiencing a wide range of temperature and precipitation. Our results support the EBF because relative brain size and the size of the optic tectum were negatively correlated with variability in temperature. In contrast, we found no association between the variability in precipitation and the length of the dry season with either brain size or the sizes of other major brain regions. We suggest that seasonality-induced food scarcity resulting from higher variability in temperature constrains brain size evolution in anurans. Less seasonal environments may therefore facilitate the evolution of larger brains in poikilothermic animals.

Muscles are vital for the process of movement, mating and escape of predators in amphibians. During evolution, the morphological and genetic characteristics as well as the size of muscles in species will change to adapt different environments. Theory predicts that low male-male competition in high-altitude/latitude selects for small limb muscles. Here, we used the Andrew’s toad (Bufo andrewsi) as a model animal to test this prediction by analyzing geographical variation in the mass of limb muscles across nine populations from the Hengduan Mountains in China. Inconsistent with the prediction, we found that latitude and altitude did not affect the relative mass of total combined limb muscles and mass of combined hindlimb muscles among populations. Meanwhile, the relative mass of combined forelimb muscles, the two forelimb muscles (flexor carpi radialis and extensor carpi radialis) and the four hindlimb muscles (e.g. biceps femoris, semimebranous, semitendinosus and peroneus) was lowest in middle latitude and largest in low latitude whereas gracilis minor was largest in high latitudes. However, we did not find any correlations between the two forelimb muscles and the four hindlimb muscles and altitude. Our findings suggest that combined forelimb muscles, flexor carpi radialis, extensor carpi radialis, biceps femoris, semimebranous, semitendinosus and peroneus are largest in low latitudes due to pressures of mate competition.

Saccharum spontaneum L. is an important wild used parent for sugarcane breeding, which can improve variety yield, ratoon ability and resistance to biotic and abiotic stress. In order to illustrate the genetic diversity of yield-related traits within a collection of S. spontaneum from Yunnan province, China, 222 clones from six geographic regions of Yunnan were screened for five yield-related traits. The results showed that: (1) the coefficient of variation (CV) of five yield-related traits ranged from 20.39 to 77.06%, of which the stalk weight exhibited the highest genetic variation with the highest CV value (77.06%), followed by the tillering rate with a CV value of 38.33%. The clones from the northwestern region of Yunnan exhibited the highest genetic variation, with an average CV value of 39.03%, followed by clones from the southwestern and southern regions; (2) the elevation and latitude of the locations from where the clones were collected had significant negative correlations with stalk height, stalk diameter and stalk weight (p

The digestive tract provides a functional relationship between energy intake and allocation. An understanding of effects of environmental factors on the evolution of digestive tract morphology is especially important. To investigate this, we studied the variation in digestive tract length across 10 populations of the Andrew’s toad (Bufo andrewsi) between 2012 and 2015 in Sichuan province, western China. These populations were collected in different habitats varying in temperature and precipitation. The results reveal an increase in the length of the digestive tract and gut with increasing temperature and decreasing precipitation, when controlling for the effect of body size. Our findings suggest that individuals of populations living in high-temperature and low-precipitation environments have longer digestive tracts, possibly because they consume less animal-based foods and more high-fiber foods.

Several previous studies have indicated that diabetic have higher risk of suffering from Alzheimer's disease, which severely induced cognitive dysfunction. However, the underlying molecular mechanism and more details on the cognitive deficits induced by hyperglycemia have not been elucidated. Here in our present study, on the basis of Goto-Kakizaki (GK) rats and streptozotocin (STZ)-induced diabetic model, we detected the variation of dendritic spine density in hippocampus as well as the differential expression of some important signal transduction molecules that were of relevance in learning and memory function. We found that the magnitude of escape latency time was significantly increased in such diabetic animals; the phosphorylated Akt/CREB; SYP and BDNF as well as other downstream molecules in hippocampus neurons were also downregulated in both diabetic groups compared to the normal groups. Thus, all of these data indicate the obstacle of neuronal pathology and the Akt/CREB signaling pathway caused by hyperglycemia that may suppress cognitive behavior, which may provide a novel way for the prevention of diabetic encephalopathy and the cognitive deficits of Alzheimer's disease.

The Expensive Brain Framework (EBF) and the Cognitive Buffer Hypothesis are commonly used to explain inter- and intraspecific variation in brain size and brain architecture. Using Andrew’s toad (Bufo andrewsi) as a model species, we investigated whether brain attributes in amphibians are shaped by the interplay between age and the length of the activity season as determined by altitude and latitude. We found significant differences in absolute and relative brain volume across 12 populations. Supporting the EBF, we observed a positive correlation between absolute and relative brain volume with season length. In particular, populations experiencing long seasons were characterized by having relatively large olfactory nerve and optic tecta. Relative, but not absolute, brain volume was also positively correlated with individual age. When accounting for the effects of body size and brain volume, however, the size of different brain structures (olfactory nerve, olfactory bulbs, telencephalon, optic tecta and cerebellum) was independent from age. Taken together, our findings are largely in agreement with the EBF to determine brain size variation in Andrew’s toad.

This study investigated effect of various durations of contton cropping on microbial community diversity of rhizosphere soil in south and north parts of Xinjiang by Biolog method,cultured-based microorganism and soil enzyme measurements.Biolog analysis showed that there were significant differences in microbial carbon utilization and functional diversities of microbial community of rhizosphere soils under various durations of contton cropping.Soil microbial activity of uncultivated soils were lower than that of cultivated soils.The AWCD of rhizosphere microbial community and Shannon(H) index were higher under 5—10 years than under 15—20 years of continuous contton cropping.The principal component analysis showed that there was significant differences in carbon utilizations of soil microbial community.The significant differences existed on PC1 and PC2,respectively,between cultivated and uncultivated soils in group one and group two,and the coefficients of uncultivated soils were negative,those of cultivated soils were positive.However significantly differences in PC1 were found between soils under rotation system and continuous cotton cropping.The differences of carbon sources in PC1 were mostly caused by carboxylic acid and polymers,which were probably the main carbon sources affectting soil microbial community under continuous cotton cropping.The results of count of soil culturable microorganism showed that numbers of bacteria in the uncultivated soils were less than that in the cultivated soils.Nevertheless,numbers of bacteria were larger under 5—10 years than under 15—20 years of continuous contton cropping.The numbers of fungi increased after 15—20 years continuous contton cropping.However,numbers of Actinomyces did not change obviously between cultivated and non-cultivated soils.Soil enzyme activities were decreased significantly after 5—15 years of continuous contton cropping,then,increased after 15—20 years of continuous contton cropping.It is concluded that soil microbial community diversities were decreased in rhizosphere soils under long-term continuous cotton cropping,and this might be the main cause of yield decrease of cotton in Xinjiang.

Five hundred and ninety-six Saccharum. spontaneum L. were selected to investigate the best sampling strategy for pre-core collection. Based on 21 quantitative and 6 qualitative traits, 50 sampling procedures were established concerning grouping principles,sampling proportions within group,selection methods within group. First grouping or non-grouping was determined based on collecting region, altitude, stalk diameter, latitude, zoology region and cluster. Then sampling within group was set up based on simple proportion, logarithmic proportion, square root proportion and genetic proportion. And random and cluster selection was applied for sampling within group. At the same time, 10 grads for sampling (5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, and 50%) from total resource were analyzed to decide the best sampling grad. The sampling strategies were evaluated according to comprehensive score based on indices of genetic diversity, variance of phenotypic value, variance of phenotypic frequency, coefficient of variation and ratio of phenotypic traits retained. The results showed that cluster sampling was better than the random sampling, collecting regions grouping and stalk diameter grouping were the best grouping and genetic proportion was the best sampling proportions within group. Finally, after analyzing 50 sampling strategies and 10 sampling grads, sampling based on stalk diameter grouping, genetic proportion and cluster sampling combined with sampling grad of 15% was confirmed to be the optimal sampling strategy for pre-core collection. A pre-core collection contained 92 samples including 5 additional ones with the rare phenotypic traits was established according to the best sampling strategy, which accounted for 15.44% of total resources. The genetic diversity, variance coefficient, variance of phenotypic frequency and variance of phenotypic value of this pre-core collection were better than that of the total resource and ratio of phenotypic traits retained came to 100%, which suggests the pre-core collection could preferably represent the total resource.

Pot culture experiments were established to determine the effects of colonization by arbuscular mycorrhizal fungi (AMF) (Glomus mosseae and G. sp) on maize (Zea mays L.) grown in Pb, Zn, and Cd complex contaminated soils. AMF and non-AMF inoculated maize were grown in sterilized substrates and subjected to different soil heavy metal (Pb, Zn, Cd) concentrations. The root and shoot biomasses of inoculated maize were significantly higher than those of non-inoculated maize. Pb, Zn, and Cd concentrations in roots were significantly higher than those in shoots in both the inoculated and non-inoculated maize, indicating the heavy metals mostly accumulated in the roots of maize. The translocation rates of Pb, Zn, and Cd from roots to shoots were not significantly difference between inoculated and non-inoculated maize. However, at high soil heavy metal concentrations, Pb, Zn, and Cd in the shoots and Pb in the roots of inoculated maize were significantly reduced by about 50% compared to the non-inoculated maize. These results indicated that AMF could promote maize growth and decrease the uptake of these heavy metals at higher soil concentrations, thus protecting their hosts from the toxicity of heavy metals in Pb, Zn, and Cd complex contaminated soils.

* These two authors contribute equally to this paper. In order to identify the proteins associated with Helicobacter pylori colonization in mice, we used 2-dimensional gel electrophoresis (2-DE) to analyze the membrane- and soluble-cellular proteins extracted from H. pylori strain 26695 and the mouse-passaged homolog 88-3887. We defined 2- and 3-fold changes in protein expression as the threshold values for differential expression in the membrane-protein and whole-cell-protein fractions, respectively. The differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF). A total of 29 proteins, including 16 mem- brane- or membrane-associated proteins (13 upregulated, 3 downregulated) and 13 cellular proteins (10 upregulated, 3 downregulated) were differentially expressed between the strains 26695 and 88-3887. Among the upregulated proteins, 10 proteins had been previously shown to be associated with the mouse colonization, and 13 upregulated proteins were shown to be associated with the adaptation of H. pylori in murine hosts for the first time in this study. The identified proteins were classified as proteins related to metabolism, stress response, virulence, or adhesion. The data presented in this report indicated that there were subsets of upregulated proteins in mouse-adapted H. pylori. In particular, the adhesins, virulence factors, and stress- response proteins are likely to contribute to colonization in mice.

Secondary metabolites not only play important ecological roles in plants but also are important pharmaceutical and source compounds for derivative synthesis. Production of plant secondary metabolites is believed to be controlled by the endogenous signal network of plants. However, the molecular basis is still largely unknown. Here we show that matrine production of Sophora flavescens Ait. cells treated with low levels of jasmonic acid (JA) and nitric oxide (NO) is significantly increased although treatment with low concentrations of JA or NO alone has no effects on matrine production, showing that JA and NO may act synergistically in triggering matrine production. Moreover, treatment with NO triggers lipoxygenase (LOX) activity and enhances JA levels of the cells, showing that NO may activate the endogenous JA biosynthesis of S. flavescens cells. External application of JA induces nitric oxide synthase-like activities and stimulates NO generation of S. flavescens cells, which suggests that JA may trigger NO generation of the cells. Thus, the results reveal a mutually amplifying reaction between JA and NO in S. flavescens cells. Furthermore, JA and NO inhibitors suppress not only the mutually amplifying reaction between JA and NO but also the synergistic effects of NO and JA on matrine production. Therefore, the data demonstrate that the synergistic action of JA and NO in inducing matrine production might be due to the mutually amplifying reaction between JA and NO in the cells.

This study was designed to investigate the effect of ghrelin on gastric acid secretion in weaning piglets both in vivo and in vitro. Thirty newborn piglets were selected from six litters and on 28, 35 (weaning), 38, 42 and 45d of age, respectively, one piglet from each litter was killed and the mucosal tissue from gastric fundus was collected for detecting ghrelin mRNA as well as H(+)-K(+)-ATPase mRNA expression and activity. Primary cultures of gastric mucosal cells from 5-week-old weaning piglets were challenged with 3x10(-5), 3x10(-4), 3x10(-3), 3x10(-2) and 3x10(-1)nmol/ml h-ghrelin, respectively, for 4h in order to further clarify the effect of ghrelin on gastric H(+)-K(+)-ATPase mRNA expression and activity. Ghrelin mRNA expression in gastric fundus kept stable from 28d to 42d, followed by a sudden increase on 45d, exhibiting a peak that was significantly higher than any other age groups investigated. H(+)-K(+)-ATPase activity and mRNA expression showed similar trends of increase with slightly different timing. H(+)-K(+)-ATPase mRNA expression tended to increase on 42d, while H(+)-K(+)-ATPase activity started to rise from 35d, but neither of them reached significantly higher levels until 45d. In vitro, ghrelin significantly increased H(+)-K(+)-ATPase activity of gastric mucosal cells at 3x10(-4), 3x10(-3), and 3x10(-2)nmol/ml, but augmented H(+)-K(+)-ATPase mRNA expression only at 3x10(-4)nmol/ml. The results indicate that ghrelin mRNA expression is up-regulated 10 days after weaning in the gastric fundus of piglets, coinciding with the increase of H(+)-K(+)-ATPase mRNA expression and activity. Ghrelin acts on gastric mucosal cells to stimulate both mRNA expression and activity of H(+)-K(+)-ATPase in vitro.

Aim: To express and purify lipoprotein-associated phospholipase A2 (Lp-PLA 2 ), and to establish a screening model for Lp-PLA 2 inhibitors using the expressed Lp-PLA 2 . Methods: We cloned the full-length cDNA of Lp-PLA 2 from differentiated THP-1 cells, and subcloned the cDNA into the baculovirus transfer vector pFastBac1. In addition, we introduced an N-terminal Kozak sequence for high-level translation initiation and a C-terminal sequence of 6 histidine residues for purification. The fusion enzyme was expressed in Sf9 insect cells and purified by Ni 2+ affinity chromatography. Recombinant Lp-PLA 2 activity was measured using [ 3 H]PAF as a substrate, and we examined the enzyme activity of recombinant Lp-PLA 2 pretreated with the known Lp-PLA 2 inhibitor SB435495. Results: We successfully cloned the full-length Lp-PLA 2 gene from differentiated THP-1 cells. The fusion enzyme was expressed in Sf9 insect cells at a high level and purified efficiently through a 2-step procedure. The recombinant Lp-PLA 2 activity was measured using [ 3 H]PAF as a substrate, and proved to be enzymatically active. Lp-PLA 2 inhibitor SB435495 produced a good inhibition curve for inhibition of recombinant Lp-PLA 2 with an IC50 of 57±1 μmol/L. Conclusion: We expressed and purified Lp-PLA 2 at a high level in insect cell-baculovirus expression system. The yield ratio was much greater than that obtained from human plasma and we established a screening model for Lp-PLA 2 inhibitors using the expressed Lp-PLA 2 .

Fungal elicitor prepared from the cell walls of Aspergillum niger induces multiple responses of Hypericum perforatum cells, including nitric oxide (NO) generation, jasmonic acid (JA) biosynthesis, and hypericin production. To determine the role of NO and JA in elicitor-induced hypericin production, we study the effects of NO scavenger 2- to 4-carboxyphenyl-4,4, 5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPITO), nitric oxide synthase inhibitor S,S′-1,3-phenylene-bis(1,2-ethanediyl)-bis-isothiourea, and inhibitors of the octadecanoid pathway on elicitor-induced NO generation, JA biosynthesis, and hypericin production. Pretreatment of the cells with cPITO and JA biosynthesis inhibitors suppresses not only the elicitor-induced NO generation and JA accumulation but also the elicitor-induced hypericin production, which suggests that both NO and JA are involved in elicitor-induced hypericin biosynthesis. S,S′-1,3-phenylene-bis(1,2-ethanediyl)-bis-isothiourea and cPITO inhibit both elicitor-induced NO generation and JA biosynthesis, while JA biosynthesis inhibitors do not affect the elicitor-induced NO generation, indicating that JA acts downstream of NO generation and that its biosynthesis is regulated by NO. External application of NO via its donor sodium nitroprusside induces hypericin production in the absence of fungal elicitor. Sodium-nitroprusside-induced hypericin production is blocked by JA biosynthesis inhibitors, showing that JA biosynthesis is essential for NO-induced hypericin production. The results demonstrate a causal relationship between elicitor-induced NO generation, JA biosynthesis, and hypericin production in H. perforatum cells and indicate a sequence of signaling events from NO to hypericin production, within which NO mediates the elicitor-induced hypericin biosynthesis at least partially via a JA-dependent signaling pathway.

Aim: To design and synthesize a series of novel amino acid-binding 1,5-diarylpyrazole derivatives, which are intended to act as prodrugs with better aqueous solubility than celecoxib, and which will exert potent anti-inflammatory activities after being converted to their parent compounds in vivo . Methods: To introduce an amino acid, celecoxib analogs containing amino or methylamino group were synthesized first through multi-step chemical reactions. All the synthesized compounds were screened in an intact cell-based assay in vivo and in carrageenan-induced mouse paw edema in vivo . Some active compounds were selected for further evaluation in a carrageenan-induced rat paw edema model. The preliminary pharmacokinetics experiments were conducted using high performance liquid chromatography/mass spectrometry (HPLC/MS). Results: Celecoxib, 6 of the 1,5-diarylpyrazole class of celecoxib analogs, and their amino acid derivatives (hydrochloride salts) were synthesized. in vivo screening, the hydrochloride salts showed decreased inhibitory effects on cyclooxygenase (COX)-1 and COX-2 compared with their parent compounds, but some exhibited potent anti-inflammatory activity in vivo . Compound 4a was selected for further evaluation, and its anti-inflammatory effect was equivalent to that of celecoxib after oral administration in the carrageenan-induced rat paw edema model. At three doses (25 mg/kg, 50 mg/kg, and 100 mg/kg) the percentage inhibition on edema was 20.7%, 52.6%, and 62.6% (for compound 4a) and 27.8%, 38.4%, and 40.1% (for celecoxib), respectively. Preliminary pharmacokinetic evaluations support the hypothesis that compound 4a was actually converted to its parent compound, compound 4. Conclusion: The compound bound with amino acid acts like prodrug, which can exert anti-inflammatory effect similar to celecoxib after being converted to its parent compound. This finding will be of great benefit in carrying out structural modifications of prodrug-like selective COX-2 inhibitors.

Sodium nitroprusside (SNP) was used as the donor of nitric oxide (NO) to investigate its effect on catharanthine synthesis and the growth of Catharanthus roseus suspension cells. The results showed that SNP at high concentrations (10.0 and 20.0 mmol/L) stimulated catharanthine formation of C. roseus cells, but inhibited growth of the cells. Low concentrations of SNP (0.1 and 0.5 mmol/L) enhanced the growth of C. roseus cells, but had no effect on catharanthine synthesis. The maximum total catharanthine production was achieved by the addition of 0.5 and 10.0 mmol/L SNP to the cultures at day 0 and day 10, respectively, being about threefold of the control. NO-induced catharanthine production of C. roseus cells was strongly suppressed by jasmonic acid (JA) biosynthesis inhibitor ibuprofen (IBU) and nordihydroguaiaretic (NDGA). The result suggests that the stimulatory role of NO on catharanthine production is partially JA-dependent.

Here we describe IS Hp609 of Helicobacter pylori , a new member of the IS 605 mobile element family that is novel and contains two genes whose functions are unknown, jhp960 and jhp961 , in addition to homologs of two other H. pylori insertion sequence (IS) element genes, orfA , which encodes a putative serine recombinase-transposase, and orfB , whose homologs in other species are also often annotated as genes that encode transposases. The complete four-gene element was found in 10 to 40% of strains obtained from Africa, India, Europe, and the Americas but in only 1% of East Asian strains. Sequence comparison of 10 representative IS Hp609 elements revealed higher levels of DNA sequence matches (99%) than those seen in normal chromosomal genes (88 to 98%) or in other IS elements (95 to 97% for IS 605 , IS 606 , and IS 607 ) from the same H. pylori populations. Sequence analysis suggested that IS Hp609 can insert at many genomic sites with its left end preferentially next to TAT, with no target specificity for its right end, and without duplicating or deleting target sequences. A deleted form of IS Hp609 , containing just jhp960 and jhp961 and 37 bp of orfA , found in reference strain J99, was at the same chromosomal site in 15 to 40% of the strains from many geographic regions but again in only 1% of the East Asian strains. The abundance and sequence homogeneity of IS Hp609 and of this nonmobile remnant suggested a recent bottleneck and then rapid spread in H. pylori populations, possibly selected by the contributions of the elements to bacterial fitness.

Elicitor prepared from the cell walls ofPenicillium citrinum induces multiple responses ofTaxus chinensis cells, including nitric oxide (NO) generation, sequentially followed by the activation of PAL and synthesis of taxol. NO scavenger cPITO and nitric oxide synthase (NOS) inhibitor PBITU prevent the latter two reactions, all of which are triggered in the absence of elicitor by NO donor sodium nitroprusside (SNP). The elicitor-induced NO release ofTaxus chinensis suspension cells is strongly inhibited by PBITU. These results demonstrate a causal relationship between NO generation and the latter two reactions ofTaxus chinensis cells to the elicitor, and also indicate that NO, produced via NOS inTaxus chinensis cells treated with fungal elicitor, might act as an essential signaling molecule for triggering the activation of PAL and synthesis of taxol.

This study aimed to investigate the protective effects of hydrogen rich water on the intestinal ischemia/reperfusion (I/R) injury in a rat intestinal intussusception (II) model. Ninety Sprague-Dawley rats were randomly assigned into three groups (n = 30 per group). In sham group, rats received laparotomy, and the intestine was exposed for 15 minutes without II. In I/R + saline group and I/R + hydrogen group, rats received II after laparotomy and then intestine was relocated 8 hours later, followed by immediately intraperitoneal injection of normal saline and hydrogen rich water (HRW) (5 mL/kg), respectively. One hour later, the intestine was collected for hematoxylin-eosin staining and immunohistochemistry for apoptotic cells and 8-oxo-deoxyguanosine, and blood was harvested for detection of tumor necrosis factor-a, malondialdehyde and superoxide dismutase. Hematoxylin-eosin staining showed the intestinal mucosa was significantly damaged in I/R + saline group, which was markedly attenuated after HRW treatment. The serum tumor necrosis factor-α content increased significantly in I/R + saline group, but HRW treatment reduced serum tumor necrosis factor-α content as compared to I/R + saline group (P < 0.05). Serum malondialdehyde content and 8-oxo-deoxyguanosine positive cells in the intestine increased dramatically after II, but HRW significantly reduced them in I/R+hydrogen group (P < 0.05). In addition, superoxide dismutase activity reduced markedly and apoptotic cells increased in I/R + saline group as compared to sham group, but they HRW increased superoxide dismutase activity and reduced apoptotic cells significantly in I/R + hydrogen group (P < 0.05). Our results indicate hydrogen rich water is able to attenuate II induced intestinal I/R injury via inhibiting intestinal inflammation, attenuating intestinal/serum oxidative stress and reducing apoptotic intestinal cells.

We report for the first time the influence of ANXA7 gene on human hepatocellular carcinoma cells (HCC). We down-regulated ANXA7 in human HCC cell line (HepG2) using siRNA method. By Western Blot analysis, we confirmed about 70% down-regulation of the gene in the shRNA-ANXA7 transfected cells (shRNA-ANXA7-HepG2) compared to the non-specific sequence shRNA transfected cells (control-shRNA-HepG2) and the un-manipulated-HepG2 cells. We used CCK-8 cell proliferation kit and observed about 65% reduction in the shRNA-ANXA7-HepG2 cells where the two controls exhibited comparable cell proliferation rates. Also, by using PI staining followed by flow cytometry, we noticed a cell cycle arrest at G0/G1 with more than one fold reduction of shRNA-ANXA7-HepG2 cell population in the S-phase of the cell cycle. Also of particular note was a significant aneuploidy in the controls compared to zero aneuploidy in the ANXA7 down-regulated cells. Migration of the cells was detected using Boyden's transwell chamber and scratch wound healing assay which showed 50% and 30% respective reductions in shRNA-ANXA7-HepG2 cells migration. Furthermore, the control-shRNA-HepG2 cells and the un-manipulated-HepG2 cells invaded through the ECM-coated transwell plates two times more than the shRNA-ANXA7-HepG2 cells. We have found ANXA7 to be functioning like a tumour promoter in HepG2 human hepatocellular carcinoma cells and could have a potential as a therapeutic window into the management of liver cancer.

Di-isopropyl phosphite reacts with bromotrifluoromethane under basic conditions giving a mixture of some seven products. Identification of these by high-resolution NMR and MS shows that 80% of all products are formed by displacement of fluorine from carbon. Dialkyl fluorophosphate is a minor product in these reactions and in additional examples involving other fluorocarbons and α-fluorophosphonates. In all cases, fluorine is removed from carbon in direct competition with either bromine or chlorine with relative halophilic ratios near unity. These processes are best under-stood in terms of fluorophilic reactions of a phosphorus anion on the C-F bond, and thereby constitute a new class of X-philic reaction.

On the basis of 103 SSR linkage map, QTLs associated with tassel branch number (TBN) and total tassel length (TTL) were studied by composite interval mapping with F3 families, which were developed from the cross N87-1 x 9526 and surveyed for phenotype under normal condition (CK) and drought-stress environment (DS). Four QTLs on chromosomes 2, 5, 7, and 10, respectively, were associated with TBN under DS, two of which were not only repeatedly detected on chromosomes 5 and 7 under the CK but also were linked to some QTLs related to drought tolerance that had already been reported in the same mapping population. The two QTLs controlling TTL were identified on chromosomes 2 and 6 under CK, while three QTLs for TTL were detected on chromosomes 2, 4, and 10 under DS. The QTL on chromosome 2 for TTL was consistent under two environments. Most of QTLs for TBN were partial additive while QTLs for TTL were dominant and over-dominant in terms of gene action.

To investigate the influence of the pyrimidine 2-keto group on selection of nucleotides for incorporation into DNA by polymerases, we have prepared two C nucleoside triphosphates that are analogues of dCTP and dTTP, namely 2-amino-5-(2'-deoxy-beta-d-ribofuranosyl)pyridine-5'-triphosphate (d*CTP) and 5-(2'-deoxy- beta-d-ribofuranosyl)-3-methyl-2-pyridone-5'-triphosphate (d*TTP) respectively. Both proved strongly inhibitory to PCR catalysed by Taq polymerase; d*TTP rather more so than d*CTP. In primer extension experiments conducted with either Taq polymerase or the Klenow fragment of Escherichia coli DNA polymerase I, both nucleotides failed to substitute for their natural pyrimidine counterparts. Neither derivative was incorporated as a chain terminator. Their capacity to inhibit DNA polymerase activity may well result from incompatibility with the correctly folded form of the polymerase enzyme needed to stabilize the transition state and catalyse phosphodiester bond formation.