Genetic diversity and population structure of the Asian green mussel Perna viridis in South China Sea based on microsatellite markers

The green mussel, Perna viridis is ecologically and economically important in the coastal region of the South China Sea. Determining its population genetic structure at this fine geographic scale will help sustainable management of natural stocks. In this study, we examined the genetic diversity and population genetic structure of P. viridis from four locations in the South China Sea (n = 45–48) using nine microsatellite loci. The results showed moderate levels of genetic diversity in all four samples (mean A = 13.222–14.000, mean A_e = 7.092–7.571, mean A_r = 12.894–13.746, mean H_o = 0.596–0.656, mean H_e = 0.690–0.733) and a large effective population size estimate for the pooled sample (total N_e estimates = infinity, 95% CI = 1869.0-infinity). We did not detect any sign of recent bottleneck events in P. viridis populations in the South China Sea. The conventional and a model-based analysis reveal low, non-significant genetic divergence among the four samples (F_ST = − 0.001–0.005, P > 0.05/6). The results obtained from this study can provide valuable genetic information for the conservation and fishery management of P. viridis by retaining the high N_e estimates.     

WEE1 inhibition and genomic instability in cancer

One of the hallmarks of cancer is genomic instability controlled by cell cycle checkpoints. The G₁ and G₂ checkpoints allow DNA damage responses, whereas the mitotic checkpoint enables correct seggregation of the sister chromosomes to prevent aneuploidy. Cancer cells often lack a functional G₁ arrest and rely on G₂ arrest for DNA damage responses. WEE1 kinase is an important regulator of the G₂ checkpoint and is overexpressed in various cancer types. Inhibition of WEE1 is a promising strategy in cancer therapy in combination with DNA-damaging agents, especially when cancer cells harbor p53 mutations, as it causes mitotic catastrophy when DNA is not repaired during G₂ arrest. Cancer cell response to WEE1 inhibition monotherapy has also been demonstrated in various types of cancer, including p53 wild-type cancers. We postulate that chromosomal instability can explain tumor response to WEE1 monotherapy. Therefore, chromosomal instability may need to be taken into account when determining the most effective strategy for the use of WEE1 inhibitors in cancer therapy.     

Genetic diversity and population structure of Buddleja crispa Bentham in the Himalaya-Hengduan Mountains region revealed by AFLP

Buddleja crispa Benth. is one of the most morphologically variable species in genus Buddleja, and it is widely distributed in the Himalaya-Hengduan Mountains (HHM) region. This study used AFLPs as a tool to examine the genetic variation among and within 25 populations of B. crispa. Analysis of population genetics of the species aimed to clarify morphological variation, current distribution patterns, strong adaptability to habitats, and the effects of geological factors in the HHM region. The genetic structure results, based on PCoA and NJ cluster analyses, revealed that the populations of B. crispa were divided into two genetic groups. Furthermore, the peripheral populations had lower genetic diversity than the populations in the center of the distribution areas (Three Parallel Rivers). We conclude that the gene flow (predominantly seed and pollen flow) and the population differentiation of B. crispa might be more affected by the barriers formed by rivers and mountains than by geographic distance.     

Genetic diversity and population differentiation of the Chinese soft-shelled turtle (Pelodiscus sinensis) in three geographical populations

The Chinese soft-shelled turtle (Pelodiscus sinensis) is one of the most important economical chelonians in the world. To understand the genetic variations of the Chinese soft-shelled turtle in China, 62 individuals were sampled from three localities and 18 polymorphic microsatellite loci tested were used to detect genetic diversity and population structure. Results showed that the genetic diversity of the wild P. sinensis was high. Except for the Wuhu populations, the majority of microsatellite loci are not deviation from Hardy–Weinberg equilibrium in the other two populations. AMOVA analysis indicated that genetic variations occurred mainly within populations (97.4%) rather than among populations (2.6%). The gene flow estimates (Nm) among three geographic populations demonstrated that strong gene flow existed (Nm > 1, mean 6). The present study supported that different habitats, breed turtles escaped, multiple paternity and long evolutionary history may be responsible for the current genetic diversity and differentiation in the wild Chinese soft-shelled turtle.     

Genetic diversity and population structure of Leptosphaeria maculans isolates in Western Canada

Leptosphaeria maculans is a serious concern for canola production worldwide. For effective disease management, knowledge of the pathogen's genetic variability and population structure is a prerequisite. In this study, whole-genome sequencing was performed for 162 of 1590 L. maculans isolates collected in the years 2007–2008 and 2012–2014 in Western Canada. DNA variants in genome-wide and specific regions including avirulence (Avr) genes were characterized. A total of 31,870 high-quality polymorphic DNA variants were used to study L. maculans genetic diversity and population structure. Cluster analysis showed that 150 isolates were clustered into 2 main groups and 4 subgroups by DNA variants located in either Avr or small secreted protein-encoding genes and into 2 main groups and 6 subgroups by genome-wide variants. The analysis of nucleotide diversity and differentiation also confirmed genetic variation within a population and among populations. Principal component analysis with genome-wide variants showed that the isolates collected in 2012–2014 were more genetically diverse than those collected in 2007–2008. Population structure analysis discovered three distinct sub-populations. Although isolates from Saskatchewan and Alberta were of similar genetic composition, Manitoba isolates were highly diverse. Genome-wide association study detected DNA variants in genes AvrLm4-7, Lema_T86300, and Lema_T86310 associated with the years of collection.     

Chromosomal instability, reproductive cell death and apoptosis induced by O-methylguanine in Mex, Mex and methylation-tolerant mismatch repair compromised cells: facts and models

O⁶-Methylguanine (O⁶-MeG) is induced in DNA by methylating environmental carcinogens and various cytostatic drugs. It is repaired by O⁶-methylguanine-DNA methyltransferase (MGMT). If not repaired prior to replication, the lesion generates gene mutations and leads to cell death, sister chromatid exchanges (SCEs), chromosomal aberrations and malignant transformation. To address the question of how O⁶-MeG is transformed into genotoxic effects, isogenic Chinese hamster cell lines either not expressing MGMT (phenotypically Mex⁻), expressing MGMT (Mex⁺) or exhibiting the tolerance phenotype (Mex⁻, methylation resistant) were compared as to their clastogenic response. Mex⁻ cells were more sensitive than Mex⁺ cells to N-methyl-N′-nitro-N-nitrosoguanidine (MNNG)-induced chromosomal breakage, with marked differences in sensitivity depending on recovery time. At early recovery time, when cells out of the first post-treatment mitosis were scored, aberration frequency was about 40% reduced in Mex⁺ as compared to Mex⁻ cells. At later stages of recovery when cells out of the second post-treatment mitosis were analyzed, the frequency of aberrations increased strongly in Mex⁻ cells whereas it dropped to nearly control level in Mex⁺ cells. From this we conclude that, in the first post-treatment replication cycle of Mex⁻ cells, only a minor part of aberrations (<40%) was due to O⁶-MeG whereas, in the second post-treatment replication cycle, the major part of aberrations (>90%) was caused by the lesion. Thus, O⁶-MeG is a potent clastogenic DNA damage that needs two DNA replication cycles in order to be transformed with high efficiency into aberrations. The same holds true for sister chromatid exchanges (SCEs). MNNG is highly potent in inducing SCEs in Mex⁻ cells in the second replication cycle after alkylation. Under these conditions, SCE induction is nearly completely prevented by the expression of MGMT. This is opposed to SCE induction in the first post-treatment replication cycle, where higher doses of MNNG were required to induce SCEs and no protective effect of MGMT was observed. This indicates that SCEs induced in the first replication cycle after alkylation are due to other lesions than O⁶-MeG. In methylation tolerant cells, which are characterized by impaired G–T mismatch binding and MSH2 expression, aberration frequency induced by MNNG was weakly reduced in the first and strongly reduced in the second post-treatment mitoses, as compared to CHO wild-type cells. The results indicate that mismatch repair of O⁶-MeG–T mispairs is decisively involved in O⁶-MeG born chromosomal instability and recombination. We also show that Mex⁺ and methylation tolerant cells are more resistant than Mex⁻ cells with regard to induction of apoptosis, indicating O⁶-MeG to be also an apoptosis-inducing lesion. The data are discussed as to the mechanism of cytotoxicity, aberration and SCE formation in cells treated with a methylating agent.     

Genetic diversity and population structure analysis of Qinghai-Tibetan plateau schizothoracine fish (Gymnocypris dobula) based on mtDNA D-loop sequences

Gymnocypris dobula is a commercially important fishery species and mostly distributed in the freshwater of Tibetan plateau. In this study, genetic diversity and intraspecific population differentiation were examined by using mitochondrial DNA D-loop sequences in 97 individuals sampled from three localities (Pali, Lasa and Yanghu) in the Tibetan plateau. Two hundred and fourteen polymorphic sites and 50 haplotypes were defined among the three localities. Genetic diversity analysis showed that the highest genetic diversity level was found in Pali population. Phylogenetic relationships analysis results indicated that closer phylogenetic relationships were found between the Yanghu and Lasa populations. Genetic population differentiation analysis indicated that the majority of variation (84.91%) was attributed to variations among populations and the largest differentiation was found between Pali and Lasa localities (F_st = 0.874, Nm = 0.036). Moreover, the historical demographic events were assessed by implementing the mismatch distribution analysis, Fajima's D test and Fu's Fs test. The results indicated that the Pali population had undergone a demographic expansion, possibly within the last 0.163 MYA (Million Years Ago). Our study firstly identified the population genetic structure of the G. dobula, which could be helpful for artificial breeding, fishery stock identification and resource conservation for this species.     

Genetic diversity and population structure of endangered Isoetes coreana in South Korea based on RAPD analysis

Isoetes coreana Chung and Choi is on the list of critically endangered species in South Korea. Using randomly amplified polymorphic DNA (RAPD) markers, we investigated the genetic diversity within and between seven local populations of I. coreana. Ten RAPD primers produced a total of 94 bands, of which 59 (62.8%) were polymorphic. A low level of genetic diversity was recognized within the populations of I. coreana: polymorphic loci (P), with values ranging from 3.4% to 33.9%, and a mean value of 15.5% being observed. Analysis of molecular variance (AMOVA) showed that genetic diversity was greater among populations (81.6%) than within populations (19.7%) or among the three regions included in this study (Han River, Youngsan River, and Nakdong River). In addition, a high degree of genetic differentiation (θ^B = 0.742) was detected among the populations. These results indicate that genetic differentiation has occurred very rapidly. However, the rate of gene flow between populations was found to be as low as 0.059, irrespective of the genetic and geographical distances between the populations, which indicates that genetic drift must have played an important role in forming the present populations of I. coreana. Because a reduction of genetic diversity as a result of genetic drift is undesirable, increasing the gene flow between populations of Korean quillwort I. coreana should be considered as a conservation strategy.     

Genetic variability and population structure of Bergenia ciliata (Saxifragaceae) in the Western Himalaya inferred from DAMD and ISSR markers

Genetic variability and population structure of Bergenia ciliata (Haw.) Sternb., commonly known as “Pashanbheda” (Stone-breaker), collected from the Western Himalayan region of India were estimated using two DNA fingerprinting methods viz., directed amplification of minisatellite DNA (DAMD) and inter simple sequence repeats (ISSR). The cumulative data analysis of DAMD and ISSR markers for 74 accessions from eight populations showed 86.1% polymorphism. Analysis of molecular variance (AMOVA) showed highest percentage of variation within individuals of populations (73.6%) and 21.7% among populations. STRUCTURE and PCoA analyses on the hierarchical partitioning of genetic diversity showed strong admixture of individuals among the eight assumed geographical populations of B. ciliata. The data suggests that high genetic flow is one of the major factors responsible for low genetic differentiation. Preservation of genetic diversity of B. ciliata is important, both to promote adaptability of the populations to changing environment as well as to preserve a large gene pool for future prospection. The present study using DAMD and ISSR markers, therefore, provide the means of rapid characterization of accessions within the populations, and thus enable the selection of appropriate accessions for further utilization in conservation and prospection programmes.     

Genetic population structure of miiuy croaker (Miichthys miiuy) in the Yellow and East China Seas base on mitochondrial COI sequences

The miiuy croaker (Miichthys miiuy) is an economically important fish species in China. However, information about population structure in this species is limited. In this study, the genetic diversity and population structure of four populations of Miichthys miiuy were investigated using a mitochondrial DNA (mtDNA) cytochrome c oxidase subunit I (COI) segment of 599 bp. A total of 35 polymorphic sites, and 31 haplotypes were defined in 109 individuals. High levels of haplotype diversity (0.731 ± 0.047) and low nucleotide diversity (0.00278 ± 0.0019) were observed in miiuy croaker populations. The results of AMOVA and pairwise fixation index (F_ST) analysis did not indicate any significant genetic differentiation among populations obtained from four sites (P > 0.05). In addition, no significant genealogical branches or clusters were recognized in the NJ tree. Analysis of neutral evolution and mismatch distribution suggested that the miiuy croaker might have experienced a recent population expansion.     

Genetic diversity and population structure of Opisthopappus longilobus and Opisthopappus taihangensis (Asteraceae) in China determined using sequence related amplified polymorphism markers

The genetic diversity and structure of four Opisthopappus longilobus and nine Opisthopappus taihangensis populations from Mts. Taihang were analyzed using SRAP markers. The results showed that O. longilobus (PPB = 95.16%, H = 0.349, I = 0.517) and O. taihangensis populations (PPB = 94.58%, H = 0.332, I = 0.504) had high genetic diversity. The genetic diversity within populations varied according to two species. Low genetic differentiation was detected among O. longilobus (Φ_ST = 0.105) and O. taihangensis (Φ_ST = 0.188) populations, respectively. Significantly genetic variation was found between O. longilobus and O. taihangensis (Φ_ST = 0.232). The dendrogram generated from pairwise genetic distance among all populations showed two distinct clusters, each corresponding to one species. A Mantel test revealed insignificant positive correlation between genetic distances and geographic distances (r_{O. longilobus} = 0.3021; r_{O. taihangensis} = 0.3658; P > 0.05). Those results indicated that self-compatibility, effective gene flow, biological traits, and historical factors may be the main factors causing geographical differentiation in the structure of O. longilobus and O. taihangensis populations.     

Genetic diversity and population structure of endangered endemic Paeonia jishanensis in China and conservation implications

Paeonia jishanensis, one of the most important ancestral species of cultivated tree peonies, is an endangered ornamental and medicinal plant endemic to China. A total of 236 individuals of P. jishanensis from 10 extant populations were analysed using 21 EST-SSR markers to assess their genetic diversity and population structure. Moderate genetic diversity levels (H_E = 0.340) and high genetic differentiation among populations (F_ST = 0.335) were revealed. Combining the results of STRUCTURE, PCoA and neighbour-joining analyses, the 10 populations were divided into four genetic groups that were significantly related to their geographical origins, which was further supported by hierarchical AMOVA showing the highest variation of 17.9% among groups. The Mantel test showed a significant positive correlation between genetic distance and geographic distance among populations (r = 0.873, P < 0.0001). The genetic structure of P. jishanensis may be due to limited gene flow hindered by vicariance and its breeding system: facultative vegetative reproduction. This study carries significant implications for the conservation and utilization of this endangered species.     

Genetic diversity and population structure of silver pomfret (Pampus argenteus) in the Indo-West Pacific revealed by mitochondrial control region sequences

Sequence variation of the mitochondrial control region in silver pomfret (Pampus argenteus) among six populations from the Indo-West Pacific region was analyzed. A total of 309 polymorphic sites were detected in the 464–466 bp segment of the control region among 165 individuals which defined 72 haplotypes. Haplotype diversity was ranged from 0.800 to 0.943 and nucleotide diversity ranged from 0.333% to 1.886%. Genetic distance and the fixation index indicated significant differences among populations in different geographical groups (P < 0.05). Median-joining network analysis data showed distinct patterns of phylogeographic structure. The present results suggested that P. argenteus populations have developed significant genetic structure, especially among geographically isolated groups.     

Genetic diversity and population structure of Taxus cuspidata in the Changbai Mountains assessed by chloroplast DNA sequences and microsatellite markers

Taxus cuspidata, a tree species with high economic value because of its anticancer properties, is experiencing severe reduction in populations across its range in China. We examined one chloroplast DNA (cpDNA) region (petA-psbE) and 9 nuclear simple sequence repeats (SSRs) loci variations among seven populations in the Changbai Mountains of China to investigate the levels of genetic diversity and population structure. A moderate level of haplotype diversity (H_T = 0.625), low nuclear microsatellite diversity (H_E = 0.261 ± 0.028), significant genetic differences (F_ST = 0.065) and substantial gene flow (Nm = 2.806) were observed. Most of the total genetic variation was partitioned within the population (87.8% and 94.0% for cpDNA and SSRs, respectively). Our haplotype identification permutation tests revealed that G_ST > N_ST, indicating an absence of phylogeographic structure in T. cuspidata. Neither STRUCTURE nor UPGMA analyses showed any geographic pattern in T. cuspidata populations. By comparatively analyzing the genetic diversity and survival situation of T. cuspidata, our results provide a theoretical foundation for the resource protection, utilization cultivation and breeding of this valuable plant.     

Observation and control in a model of a cell population affected by radiation

The effect of radiation on a cell population is described by a two-dimensional nonlinear system of differential equations. If the radiation rate is not too high, the system is known to have an asymptotically stable equilibrium. First, for the monitoring of this effect, the concept of observability is applied. For the case when the total number of cells is observed, without distinction between healthy and affected cells, a so-called observer system is constructed, which, at least near the equilibrium state, makes it possible to recover the dynamics of both the healthy and the affected cells, from the observation of the total number of cells without distinction.     

Genetic diversity and population structure of Armillaria luteo-virens (Physalacriaceae) in Qinghai-Tibet Plateau revealed by SSR markers

Armillaria luteo-virens is a type of ectomycorrhiza whose fruit body is widely used as an edible mushroom in the Qinghai-Tibetan Plateau. In total, 404 individuals of A. luteo-virens from 23 geographically separate populations were analyzed using eight pairs of SSR markers. The average number of alleles among nine populations was 28.65 with a range from 20 to 38. The mean observed (H_O) and expected (H_E) heterozygosity ranged from 0.446 to 0.501 and from 0.499 to 0.554, respectively. An analysis of molecular variance (AMOVA) showed that most genetic variation occurred within populations (83.32%). The mean value of F_ST (0.176) suggested low genetic differentiation among populations. The 23 studied populations were divided into two main clusters based on the NJ dendrogram, which was also confirmed by STRUCTURE analysis. This study may aid in not only the protection of this species but also the management of genetic resources for future artificial cultivation.     

Genetic diversity and population structure of Schizopygopsis younghusbandi Regan in the Yarlung Tsangpo River inferred from mitochondrial DNA sequence analysis

The genetic diversity and population structure of Schizopygopsis younghusbandi from six sites in the middle reach of the Yarlung Tsangpo River were examined based on mitochondrial DNA cytochrome b (Cyt b) gene and control region (D-loop) sequences. 1141 bp of complete Cyt b sequences and 737 bp of partial D-loop sequences for 153 individuals of S. younghusbandi were obtained by using PCR amplification and sequencing. The results showed that S. younghusbandi populations had high haplotype diversity and low nucleotide diversity, and the population genetic diversity of this species was at a low level. Analysis of molecular variance revealed that most genetic variation occurred within populations, and that genetic differentiation was at low or moderate levels. The network of haplotypes based on Cyt b gene showed that there were two groups within the examined populations. Neutrality tests and mismatch distribution analysis suggested that this species might have undergone a population expansion and the expansion time was estimated to be 0.25–0.46 Ma BP. All these results would be crucial to establish scientific strategies for effective conservation and sustainable exploitation of wild resources of S. younghusbandi.     

Cell line studies and analytical measurements of three paclitaxel complex variations

The copper(II) cation, sucrose, and hydroxychloroquine were complexed with the chemotherapy agent paclitaxel and studied for medicinal activity. Data (GI₅₀, LD₅₀) from single dose and five dose National Cancer Institute sixty cell line panels are presented. Analytical measurements of different complexes were made using Nuclear Magnetic Resonance (¹H NMR), Matrix Assisted Laser Desorption Ionization-Time of Flight-Mass Spectrometry (MALDI-TOF-MS) and Fourier Transform-Ion Cyclotron Resonance (FT-ICR). Molecular modeling is utilized to better understand the impact that species could have on physical parameters associated with Lipinski’s Rule of Five, such as log P and TPSA. On average, Cu(II) and hydroxychloroquine decreased GI₅₀ values, while sucrose increased GI₅₀ values of paclitaxel.     

一株新的人肾癌原代细胞系的建立及其生物学特性鉴定

目的:取肾癌病人标本建立一株新的人肾癌细胞系,初步对该细胞系进行鉴定,为进一步的肾癌基础研究提供实验模型.方法:2008-2009年间共采集20例肾癌新鲜手术标本,于手术后1 h内将每例标本切取四块0.5cm× 0.5 cm× 0.5 cm组织块,分别包埋于2只裸鼠的右后肢皮下及背部皮下,连续传代3次,取移植瘤体外培养,记录细胞株的生长曲线,测定细胞集落形成率,对细胞进行DNA含量测定,进行染色体分析及病理学检查.结果:其中1只裸鼠皮下移植瘤生长,继续传代,肿瘤生长速度明显加快.取移植瘤标本体外培养得到肾癌细胞系XJG-9201.形态结构,分化程度与原发瘤一致,染色体众数为65.细胞群体倍增时间为38.2h,细胞周期分析G1期62.7%,G2期11.2%,S1期25.3%,集落形成率为70%.结论:肾癌细胞系XJG-9201与原发癌保持相同的生物学特性,体外连续传代1年以上传115代.细胞形态不变,生长周期恒定,已成为一个稳定的细胞系.