蛋白质序列间进化距离的估计

本文考察了目前采用的估计同源蛋白质序列间进化距离的方法的缺陷，并提出了几个新的计算公式，它们考虑了氨基酸位点间显然存在的替代速率的差异。另外，提出了一种考虑氨基酸间不同替代概率的最大似然估计方法。文中对这些公式进行了计算比较。

4种作物果糖激酶基因的鉴定与比较分析

【目的】基于全基因组序列,鉴定与分析水稻、玉米、高粱和谷子4种作物的果糖激酶(fructokinase, FRK)家族基因,明确其复制起源方式、系统发生关系、序列及表达分化特征等,为深入开展FRK基因功能研究、改良作物产量和品质等提供数据。【方法】采用基因组搜索、基因组内及组间比较等多种方法,分析4种作物FRK家族成员的基因结构、保守基序、系统进化、复制扩张和基因表达分化等。【结果】鉴定到水稻13个、玉米16个、高粱14个、谷子16个FRK基因家族成员,其中水稻11个、玉米7个、高粱12个、谷子13个基因为离散复制基因,水稻、高粱和谷子各有1对ρWGD基因,玉米有8个WGD基因(1个ρWGD基因,7个mWGD基因)。该家族成员亚家族间基因结构差异大,但在亚家族内具有保守性。基因表达模式分析发现,直系同源基因间的表达模式相似,ρWGD基因对发生了明显的表达分化,而玉米物种特异的mWGD基因对的表达差异较小。【结论】探明了4种作物基因组包含的FRK家族基因,揭示了其复制起源、基因结构、序列及表达模式的演化特征。

荠菜属CaCA家族基因拷贝数变异与表达分化的研究

荠菜属植物是十字花科中分布较广泛的植物,其中四倍体荠菜(Capsella bursa-pastoris)是一种由祖先二倍体亲本杂交再经多倍化后形成的多倍体物种,在地域分布及多样化的逆境适应能力方面与其祖先亲本相比具有明显优势,目前四倍体荠菜对于环境适应性的研究报道还不多见.植物钙/阳离子逆转运蛋白超家族(CaCAs)是一类膜定位的离子转运蛋白,在植物抗冷、抗盐等信号通路中起到重要作用.本研究通过生物信息学及表达谱分析手段,发现四倍体荠菜CaCAs的13对亚基因组基因中有4对出现了拷贝数变异,其中3对基因在其编码的氨基酸序列和蛋白质跨膜结构方面与其直系同源基因间存在较大差异,且在冷响应表达上与拟南芥相比出现了明显偏倚.此外,与四倍体荠菜的祖先亲本Capsella rubella相比,四倍体荠菜CaCAs基因的组织表达分化较为普遍.CaCAs基因在四倍体荠菜中的拷贝数变异和表达分化现象提示了CaCAs对植物环境适应性的贡献.

Sequence Difference of Mitochondrial DNA Control Region and Genetic Differentiation of Cervinae in China

[Objective] The study aimed at analyzing Genetic Differentiation of Cervinae. [Method] The complete mitochondrial DNA control region of five species of Cervinae was determined by direct DNA sequencing. Homologous sequences of others Cervinae were gained from GenBank, and then analyzed through biology software. [Result] The lengths of their control region were 921-1 072 bp, the nucleotide content of T, A, C, and G was 32.1%, 30.2%, 22.7% and 15.0% separately. Genetic distance of each species ranged from 0.062-0.106, which belonged to level of inter-genus. The molecular phylogenetic tree indicated that red deer, skia deer and White-lipped Deer had close relationship, and they constituted one branch with sambar deer. Eld＇s deer and elk deer constituted the second branch, and hog deer constituted the third branch alone. [Conclusion] Hog deer and fallow deer should be incorporated in Cervus, divergence time is about 1.55-2.60 million years among Cervinae in China.

Segmental Duplications Are Common in Rice Genome

Segmental duplications on rice (Oryza sativa L.) chromosomes 8, 9, 11, and 12 were studied by examining the distributions of sequences resolved by 13 probes detecting multiple copies of DNA sequences. Four of the hybridization bands detected by a repetitive sequence probe, rTRS, were mapped to the ends of all the four chromosomes. Two or three of the bands detected by each of the other 12 probes were also mapped to different chromosomes. The bands detected by the same probe usually occurred in similar locations of different chromosomes. Loci detected by different DNA probes were often similarly arranged on different chromosomes. Chromosomes 8 and 9 showed colinearity of marker loci arrangement indicating a possible common origin. A segment on chromosome 9 was also very similar to the previously reported duplicated fragments on the ends of chromosomes 11 and 12 which were also detected in this study, indicating a likely common origin. Moreover, the various degrees of distributional similarity of the segments suggest a complex relationship among the chromosomes in the evolution of the rice genome. These results support the proposition that chromosome duplication and diversification may be a mechanism for the origin and evolution of the chromosomes in the rice genome.

Sequence and Molecular Evolution Analysis of Ubiquitin Proteins Encoded by Baculoviruses

[Objective] The aim of this study was to analyze the sequence characteristics and molecular evolution of ubiquitins encoded by baculoviruses.[Methods]Clustal W software was used for multiple sequence alignment analysis,and neighbor-joining method（NJ）and maximum parsimony method（MP）were used for the construction of phylogenetic tree.[Results]The baculoviral ubiquitins showed 73%-86% sequence identity to eukaryotic ubiquitin.Two heterogeneous regions of baculoviral ubiquitins were observed：one was the residues from 15-32,the other was located from residues 53 to 60.The else parts were conserved,where many functional amino acids were also observed.Phylogenetic analysis indicated that baculoviral ubiquitins could be divided into three sub-families,including sub-family GV,sub-family I and sub-family II.The molecular evolution of baculoviral ubiquitins might be under negative selection to maintain their functional and structural stability.[Conclusion]The analysis had provided reference for the researches on functional characterization of baculoviral ubiquitins.

Molecular Characterization of Four ADF Genes Differentially Expressed in Cotton

Actin depolymerizing factor （ADF）, highly conserved in all eukaryotic cells, is a low molecular mass of actin-binding protein, which plays a key role in modulating the polymerizing and depolymerizing of the actin filaments. Four cDNAs （designated GhADF2, GhADF3, GhADF4, and GhADF5, respectively） encoding ADF proteins were isolated from cotton （Gossypium hirsutum） fiber cDNA library. GhADF2 cDNA is 705 bp in length and deduces a protein with 139 amino acids. GhADF3 cDNA is 819 bp in length and encodes a protein of 139 amino acids. GhADF4 cDNA is 804 bp in length and deduces a protein with 143 amino acids. GhADF5 cDNA is 644 bp in length and encodes a protein of 141 amino acids. The molecular evolutionary relationship of these genes was analyzed by means of bioinformatics. GhADF2 is closely related to GhADF3 （99% identity） and PetADF2 （89% identity）. GhADF4 is closely related to AtADF6 （78% identity）, and GhADF5 is closely related to AtADF5 （83% identity）. These results demonstrated that the plant ADF genes are highly conserved in structure. RT-PCR analysis showed that GhADF2 is predominantly expressed in fiber, whereas, GhADF5 is mainly expressed in cotyledons. On the other hand, it seems that GhADF3 and GhADF4 have no tissue specificity. Expression levels of different ADF genes may vary considerably in the same cell type, suggesting that they might be involved in regulating tissue development of cotton and the each ADF isoform may diverge to form the functional difference from the other ADFs during evolution.

Characterization and expression of sweetfish （Plecoglossus altivelis） cathepsin D

Cathepsin D （CTSD） is a lysosomal acidic endoproteinase that plays an important role in immune response, In this study, we obtained sweetfish （Plecoglossus altivelis） CTSD （PaCTSD） via de-novo transcriptome sequencing of sweetfish macrophages. The full length cDNA sequence of PaCTSD was 1955 bp encoding a propeptide of 397 amino acids. The deduced protein had a calculated molecular weight of 43.17x 103. Multiple alignment with other known CTSD amino acid sequences revealed amino acid conservation through the teleosts. Phylogenetic tree analysis showed that PaCTSD grouped tightly with other fish CTSD, and was close to that of Atlantic salmon and rainbow trout. Subsequently, PaCTSD was prokaryotically expressed and refolded by the urea gradient method on a nickel-nitrilotriacetic acid column. Enzyme activity analysis showed that PaCTSD exhibited pH-dependent proteolytic activity. Quantitative real-time PCR showed that PaCTSD mRNA was expressed in all detected tissues in healthy sweetfish. The highest expression was observed in the spleen and white blood cells, followed by liver, head-kidney, kidney, intestine, gill, and muscle. After Listonella anguillarum infection, PaCTSD transcripts were up-regulated significantly in liver, spleen, white blood cells, and head-kidney of sweetfish. In summary, PaCTSD has proteolytic activity and is closely involved in the immune response of sweetfish.

SEQUENCE CONSERVATION OF HUMAN CYTOMEGALOVIRUS UL140 OPEN READING FRAME IN CLINICAL STRAINS

Objective To investigate the variability of human cytomegalovirus （HCMV） UL140 open reading flame （ORF） in clinical strains, and to explore the relationship between the variability of UL140 ORF and different symptoms of HC-MV infection. Methods HCMV UL140 ORF was amplified by polymerase chain reaction and sequenced selectedly in 30 clinical strains. Results UL140 ORF of all clinical strains was amplified successfully. Compared with that of Toledo strain, the nucleotide and amino acid sequence identities among all strains were 96.5% -100.0% and 95.2% -100. 0%, respectively. All of the nucleotide changes were substitutions. The post-translational modification sites were conserved. The result of phylogenetic tree showed that the strains did not cluster according to different clinical symptoms. Conclusion HCMV UL140 ORF in clinical strains is highly conserved, which may play an important role in HC-MV infection.

Runoff Change of Naoli River in Northeast China in 1955–2009 and Its Influencing Factors

Runoff change and trend of the Naoli River Basin were studied through the time series analysis using the data from the hydrological and meteorological stations. Time series of hydrological data were from 1957 to 2009 for Bao′an station, from 1955 to 2009 for Baoqing station, from 1956 to 2009 for Caizuizi station and from 1978 to 2009 for Hongqiling station. The influences of climate change and human activities on runoff change were investigated, and the causes of hydrological regime change were revealed. The seasonal runoff distribution of the Naoli River was extremely uneven, and the annual change was great. Overall, the annual runoff showed a significant decreasing trend. The annual runoff of Bao′an, Baoqing, and Caizuizi stations in 2009 decreased by 64.1%, 76.3%, and 84.3%, respectively, compared with their beginning data recorded. The wet and dry years of the Naoli River have changed in the study period. The frequency of wet year occurrence decreased and lasted longer, whereas that of dry year occurrence increased. The frequency of dry year occurrence increased from 25.0%-27.8% to 83.9%-87.5%. The years before the 1970s were mostly wet, whereas those after the 1970s were mostly dry. Precipitation reduction and land use changes contributed to the decrease in annual runoff. Rising temperature and water project construction have also contributed important effects on the runoff change of the Naoli River.

Genomic Signal Enhancement by Clustering

Weight matrix models for signal sequence motif are simple. A main limitation of the models is the assumption of independence between positions. Signal enhancement is achieved by taking the total likelihood as the objective function for maximization to cluster sequences into groups with different patterns. As an example, the initial and terminal signals for translation in rice genome are examined.

Sequence Analysis of Attachment Gene of Lumpy Skin Disease and Sheep Poxviruses

In Egypt,protection of cattle against lumpy skin disease (LSD) was carried out using a sheep poxvirus (Kenyan strain) vaccination strategy.In the present study 15 skin nodules from LSD suspected cows and 5 scab samples from sheep pox (SP) suspected sheep were collected.Hyperimmune rabbit sera to Lumpy skin disease virus (LSDV)/Ismailyia88 strain and sheep pox virus (SPV)/ Kenyan vaccinal strain were prepared.The causative agent in the collected samples was identified using immunoflourescence (IF) and immunoperoxidase techniques.Of the 15 skin nodules suspected of LSD,10 showed a positive reaction and 3 out of 5 skin scabs suspected of sheeppox were found to be positive.An antigenic correlation between field skin isolate of LSDV,tissue culture adapted LSDV/Ismailyia88 strain,field skin isolate of SPV and SPV/Kenyan vaccinal strain was studied using prepared hyperimmune sera.Also,nucleotide sequence of the PCR amplified attachment gene fragments of field skin isolate of LSDV,tissue culture adapted LSDV/Ismailyia88 strain,field skin isolate of SPV and SPV /Kenyan vaccinal strain were compared.The results revealed that the four used viruses were antigenically identical.Sequence analysis indicated that field skin LSDV isolate is more related to tissue culture adapted LSDV/Ismailyia88 strain than to vaccinal SPV/ Kenyan strain and the skin isolate of SPV is more closely related to field skin isolate of LSDV than to SPV/Kenyan vaccinal strain.Thus,further study should be applied on the advantage of a LSD vaccine prepared from LSDV in protection of cattle against LSD compared to the commonly used sheep pox vaccine.

Change Point Detection and Trend Analysis for Time Series

Trend analysis and change point detection in a time series are frequent analysis tools.Change point detection is the identification of abrupt variation in the process behaviour due to natural or artificial changes,whereas trend can be defined as estimation of gradual departure from past norms.We analyze the time series data in the presence of trend,using Cox-Stuart methods together with the change point algorithms.We applied the methods to the nearsurface wind speed time series for Australia as an example.The trends in near-surface wind speeds for Australia have been investigated based upon our newly developed wind speed datasets,which were constructed by blending observational data collected at various heights using local surface roughness information.The trend in wind speed at 10 m is generally increasing while at 2 m it tends to be decreasing.Significance testing,change point analysis and manual inspection of records indicate several factors may be contributing to the discrepancy,such as systematic biases accompanying instrument changes,random data errors(e.g.accumulation day error)and data sampling issues.Homogenization technique and multiple-period trend analysis based upon change point detections have thus been employed to clarify the source of the inconsistencies in wind speed trends.

Molecular identification and culture trials of Eutreptiella gymnastica (Eutreptiales, Euglenophyceae)

During a survey for marine microalgal resources, we isolated a rare marine euglenoid from the coastal waters of Qingdao, China in 2009, and established a pure culture. Electron microscopic and molecular phylogenetic （18S rDNA and 16S rDNA sequences） analyses revealed a close affinity with Eutreptiella gymnastica, a bloom-forming species. Different culture conditions were monitored to understand optimal E. gymnastica growth characteristics. The optimal growth conditions in a batch culture of this isolate were 20C, 160 gmol photons/（mE.s） of white light, and a salinity of 10-31. Nutrient experiments demonstrated that growth increased dramatically with a phosphorus concentration greater than 72 gmol/L. Understanding the effect of culture conditions on E. gymnastica may help understanding the blooming mechanism of this alga in its natural environment.

Isolation and Purification of Carp Mitochondrial DNA and Structural Analysis of Its tRNA^(Cys) Gene and the Light Strand Origin

A new method is presented with which we isolated milochondrial DNA from fresh carp liver usingdifferential centrifugation and DNase treatment that gave high yield of purified product with an easyand economical procedure. Highly distinct bands were displayed in agarose gel electrophoresls ofthe product digested with restrictlon enzymes, which were successfully used in constructingrestriction map and molecular clone of mitochondrial genes. With DNAs thus obtained, we havecloned cysteine tRNA gene (tRNA^(Cys) gene) of carp mitochondria, determined the nucleotide sequenceof it and the light strand origin, and depicted the cloverleaf secondary structure of tDNA^(Cya) and thelight strand origin. Analysis of nucleotide sequences of tRNA^(Cy) genes of 5 vertebrates has revealedunusual features of carp mitochondrial tRNA^(Cy) gene as compared with their cytoplasmic counter-parts, Altogether 36 bases were found in the light strand origin of carp mitochondriaf: 11 pairs in thestem; and 14 bases in the loop. As compared with those of other 11 vertebrate species, the sequenceof the stem is very conservative while both sequence and length of the loop are quite variable. Thestructure of the stem-loop may play an important role in light strand replication.

Variations of Terrestrial Water Storage in the Yangtze River Basin under Climate Change Scenarios

In this study, the water balance-based Precipitation-Evapotranspiration-Runoff (PER) method combined with the land surface model Variable Infiltration Capacity (VIC) was used to estimate the spatiotemporal variations of terrestrial water storage (TWS) for two periods, 1982-2005 (baseline) and 2071-2100, under future climate scenarios A2 and B2 in the Yangtze River basin. The results show that the estimated TWS during the baseline period and under the two future climate scenarios have similar seasonal amplitudes of 60-70 mm. The higher values of TWS appear in June during the baseline period and under the B2 scenario, whereas the TWS under A2 shows two peaks in response to the related precipitation pattern. It also shows that the TWS is recharged from February to June during the baseline period, but it is replenished from March to June under the A2 and B2 scenarios. An analysis of the standard derivation of seasonal and interannual TWS time series under the three scenarios demonstrates that the seasonal TWS of the southeastern part of the Yangtze River basin varies remarkably and that the southeastern and central parts of the basin have higher variations in interannual TWS. With respect to the first mode of the Empirical Orthogonal Function (EOF), the inverse-phase change in seasonal TWS mainly appears across the Guizhou-Sichuan-Shaanxi belt, and the entire basin generally represents a synchronous change in interannual TWS. As a whole, the TWS under A2 presents a larger seasonal variation whereas that under B2 displays a greater interannual variation. These results imply that climate change could trigger severe disasters in the southeastern and central parts of the basin.

Effect of Long-Term in Vitro Sub-culturing on Quality Degeneration of Sweet Potato Varieties： Morpho- Anatomic Assessment and Simple Sequence Repeats Analysis

The aim of this study was to assess the effect of long-term in vitro sub-culturing on the varietal degeneration of three sweet potato varieties, namely, Monate, Mokone and Ndou which were sub-cultured for 32, 23 and 12 generations, respectively. Each generation was cultured in a media which is made from 4.43 g/L Murashige and Skoog （MS）, 30 g/L sucrose and 2 g/L gelrite, respectively, and grown under 16 h light and 8 h dark photoperiod for 30 d. For each generation, 45 plantlets were acclimatized for two months in a glasshouse. Data on in vitro growth performance and 11 morphological characteristics during acclimatization were recorded. Early root and shoot formation was observed after the 27th and 21st sub-cultured generations of Monate and Mokone, respectively. During acclimatization, plantlets from the same variety showed differences in morphological traits such as leaf colour, abaxial leaf pigmentation, vine pigmentation, petiole pigmentation, leaf wrinkling and flowering. However, the rate of these morphological differences is random and irrespective to increase in sub-culturing. Therefore, to understand the genetic base of these morphological variability, two plantlets from each variety were subjected to genetic analysis by using five simple sequence repeat （SSR） primers （IB-242, IB-318, IB-255F, 1B-248 and IB-255）. Although SSR loci IB-255F and IB-318 could distinguish between the three varieties, there were no allelic polymorphisms detected in plantlets from the same varieties. Therefore, long-term sub-culturing do not leads to quality degeneration in the three sweet potato varieties.

Evaluation of Materials for the Separation of Germanium from Gallium, Zinc and Cobalt

In the past ^68Ge （Germanium-68） was purified with toxic organic solvents in liquid-liquid extractions making the product unacceptable by the FDA （food and drug Administration） for human use. The authors report initial studies utilizing three or four columns consisting of sequences of AG 1, AG50, Chelex 100 and/or Sephadex G25 resins. Five purification methods were examined and a separation consisting of the sequence： AG 1-Chelex100-Sephedex G25 provided 87% recovery of germanium. Fractionation of the elution resulted in high germanium concentrations.

Identification of Cytochrome P450 (CYP) Genes in Zhikong Scallop (Chlamys farreri)

Cytochrome P450 （CYP） superfamily is one of the membership largest and function most diverse protein superfamily recogniozed among living beings. Members of this superfamily were further assigned to different families and subfamilies based on their amino acid similarities. According to their phylogenetic relationships, the CYP genes which likely diverged from common ancestor gene and may share common functions were grouped into one clan. Widely distributing scallops are a group of the most conspicuous bivalve; however the studies on their CYP is acarce. In this study, we searched the genome and expressed sequence tags of Zhikong scallop （Chlamysfarreri） for CYP genes. In total, 88 non-redundant CIfP were identified, which were homed in 13 CYPs gene families. Phylogenetic analysis divided these genes into 4 CYP clans. As in deuterostomes, Clan 2 was the largest, which contained 33 genes belonging to CYP1, CYP2, CYP17 and CYP356 families. Clan 3 contgained 19 genes belonging to CYP3, CYP5 and CYP30 families. Clan 4 contained 23 genes, all belonging to CYP4 family. The mitochondrial CYP clan contained 9 genes belonging to CYPIO and CYP24 families. In comparison, protostomes （C. farreri, D. pluex, D. melanogaster） contained more CYP genes than deuterostomes （S. purpuratus and vertebrates） in Clan 2 but less genes in Clan 3 and Clan 4. Our findings will aid to deciphering CYP function and evolution in scallops and bivalves.

Cloning and Bioinformatics Analysis of zmERECTA

[Objective] This study was conducted to clone zmERECTA gene according to Arabidopsis ERECTA sequence and predict its characteristics by bioinformatics. [Method] The c DNA of zmERECTA gene was isolated from B73 using RT-PCR, and analyzed by bioinformatics methods. [Result] zmERECTA gene was 2 985 bp in size, which encoded a protein consisting of 944 amino acids, containing leucine-rich repeats, a PKC domain, two transmembrane regions, 14 N-glycosylation potential sites and41 kinase specific phosphorylation sites. The theoretical p I and molecular weight of zmERECTA protein was 6.01 and 10 8495.5respectively. [Conclusion] Cloning and bioinformatics of zmERECTA gene laid a foundation for further research.