抗坏血酸、谷胱甘肽参与的信号转导途径在提高植物食品营养品质中的作用

ASA(抗坏血酸)、GSH(还原型谷胱甘肽)是植物体内一类低分子量、可溶性的抗氧化剂。在动植物体内,GSH合成途径大致相同而ASA合成途径却差别很大。通过调控与二者合成有关的酶进而提高它们在植物食品中的含量具有很大的应用前景。然而,众多的环境因素,尤其是氧化胁迫通过一定的信号转导途径对植物体内与ASA、GSH合成相关酶的影响很大,酶活性的变化又影响了体内ASA、GSH含量。因此深入研究植物体内GSH、ASA参与的信号转导途径对提高植物食品营养品质具有重要意义。

Genome-wide Analysis of Glucose-6-phosphate Dehydrogenase(G6PDH) and Its Evolution in Eucalyptus grandsis

[Objective] The aim of this study was to perform genome-wide analysis of glucose-6-phosphate dehydrogenase（G6PDH） and reveal its evolution in Eucalyptus grandsis.[Method] The gene character,protein sequence and phylogenetic tree of G6PDH gene were analyzed by BLAST and other bioinformatics software within Eucalyptus grandsis whole genome database.[Result] Six G6PDH genes,including one cytomic type and five plastids,were detected in the E.grandsis genome.All the G6PDHs have conserved motifs of motif 1,motif 2,motif 3,motif 7,motif 9 and motif 11.Furthermore,promoter sequences of all E.grandsis G6PDH contain TATA box,enhancer,light-responsive,hormone-responsive and stress-responsive regulatory elements.[Conclusion] This study provided reference for the further revealing molecular function of E.grandsis G6PDH gene family

Molecular Composition and Evolution of the Chalcone Synthase (CHS) Gene Family in Five Species of Camellia (Theaceae)

The molecular composition and evolution of the chalcone synthase (CHS) gene family from five species in Camellia (Theaceae) are explored in this study. Sixteen CHS exon 2 from four Camellia species were amplified from total DNA by PCR method. Three sequences of the fifth species in Camellia and two sequences of Glycine max as the designated outgroups were obtained from GenBank. Our results indicated that CHS gene family in Camellia was differentiated to three subfamilies (A, B, C) during the evolutionary history with six groups (A1, A2, A3, BI, B2, C). Among them, only group A2 was possessed by all five species in this study. However, the other five groups were detected only in some species of the plants studied. All members of CHS gene family in this study had high sequence similarity, more than 90% among the members in the same subfamily and more than 78% among different subfamilies at nucleotide level., According to the estimated components of amino acids, the function of CHS genes in Camellia had been diverged. The nucleotide substitutions of the different groups were not identical. Based on phylogenetic analyse inferred from sequences of CHS genes and their deduced amino acid sequences, we concluded that the CHS genes with new function in this genus were evolved either by mutations on several important sites or by accumulation of the mutations after the gene duplication. A further analysis showed that the diversification of CHS genes in Camellia still occurred recently, and the evolutionary models were different to some extant among different species. So we assumed that the different evolutionary models resulted from the impacts of variable environmental elements after the events of speciation.

Progress in the Breeding of Soybean for High Photosynthetic Efficiency

Studies for many years have indicated that the seed yield of (Glycine max L. Merr.) soybean can be increased by increasing photosynthetic efficiency. The yield of cultivars with high photosynthetic efficiency (HPE) increased by 30% - 40% in comparison with the cultivars with normal photosynthetic efficiency, indicating that the breeding of soybean by increasing RPE may have a bright prospect. HPE breeding can be used as the temporal monitoring in the breeding process to avoid the divergency of the predetermined goal, although HPE breeding does not shorten the breeding time. It was observed that limited C-4 pathway exists in soybean leaf and pod, suggesting that by increasing the genetic expression of some C-4 enzymes in C-3 crops through traditional or genetic engineering techniques, new breakthroughs in increasing the photosynthetic efficiency of C-3 plant may be practicable in the future.

A Biosystematic Study on Asplenium sarelii Complex

The series Variantia Ching et S. H. Wu mainly occur in China and its members are highly variable in morphology. The denomination on this group of Asplenium is very confused in the herbaria. We hop e by means of a biosystematic study to find out their genetic relationships in the reticulate evolution, and to raise a suggestion on their taxonomic treatment. Evidence from cytology, allozyme, morphology, and palynology shows that three ancestor diploids have formed Asplenium sarelii complex comprising 13 members. A. sarelii Hook. should be typified as a diploid. The so-called tetraploid 'A. sarelii' before is an allotetraploid that comes from the doubled hybrid between diploid A. sarelii and A. tenuicaule Hayata, which should be treated as a new species A. wudangense Z. R. Wang et X. Hou. A. pekinense Hance is an autotetraploid that comes from the doubled diploid ancestor A. sarelii. A. lushanense C. Chr., a diploid species and the only ancestor of A. yunnariense group, should not been sunk as a synonym of tetraploid A. yunnariense Franch. Most probably, A. varians Wall. ex Hook. et Grev. is an autotetraploid of A. tenuicaule Hayata. Three new natural tetraploid hybrids and their origins have been found out: they are A. x longmenense ( = A. pekinense x varians), A. x jingyunense ( = A. pekinense x yunnanense) and A. x kidoi ( = A. pekinense x wudangense). Three other new natural triploid hybrids have been found and their origins have been inferred: they are A. X huawuense ( = A. sarelii X wudangense), A. x luyunense ( = A. lushanense x yunnanense) and A. x teniuvaians ( = A. tenuicaule x varians). The method of allozyme comparion combined with cytological observation is employed to reveal the complicated relationships among the members of Asplenium sarelii complex in reticulate evolution and proved to be a highly effective tool to investigate the origin of polyploid and hybrid.

AN ALLOZYME ELECTROPHORESIS STUDY ON ELEVEN SPECIES OF MEGOPHRYINAE IN CHINA

Allozymes of eleven species of Megophryinae in China were examined electrophoretically to investigate genetic diversity and phylogenetic relationships. Fourteen enzymes, presumptively coded by 24 loci were detected to be variable. Gene frequencies of each population at each locus were presented. The commonly used measure of genetic diversity, the average heterozygosity (H) were calculated based on gene frequencies. The results indicated that Megophryinae had a high level of genetic diversity in amphibians, an average H of 0.18, ranging from 0.058 to 0.28. Nei's (1978) genetic distances(Nei's D) were calculated for all possible population pairs. A dendrogram of 13 populations representing 11 species, 3 genera of Megophryinae were derived and presented by using UPGMA, based on Nei' s D. The assignment of Ophryophryne as a distinct genus were supported by an average Nei's D of 1.4067 which separated O. microstoma from all other populations.Subdivision of Brachytarsophrys from Megophrys was not supported by this study. Within Megophrys, three groups were recognized: (1)M. lateralis, M. giganticus and M. longipes; (2)M. palpebralespineosa, M. boettgeri and M. parva;(3) M. minor and M. kuatunensis. Three populations of M. omeimontis were closely related and share a clade independent from all other Megophrys, and B. feae as well.

Cloning and Phylogenetic Analysis of Ubiquitinconjugating Enzyme Gene TaUBC4 in Different Wheat Cultivars

[Objective] This study aimed to clone ubiquitin-conjugating enzyme gene TaUBC4 from different wheat cultivars and thus analyze their phylogenetic relationship.[Method] The UBC4 coding sequences were cloned through reverse transcription PCR （RT-PCR） from 21 wheat varieties.After sequencing,the UBC4 sequence in wheat cultivar Zhongguochun （GenBank accession No：M28059） was selected as the reference gene,to analyze the mutation frequency and evolutionary distance in the CDSs and corresponding amino acid sequences of the different wheat cultivars.Moreover,the phylogenetic tree based on the amino acid sequences of these TaUBC4 genes were constructed,involving the homologous sequences of TaUBC4 in eight other monocots.[Result] TaUBC4 sequence was highly conserved because the similarity in DNA sequences of the wheat varieties was over 94％,while that in amino acid sequence was over 96％.And the amino acid sequence difference only can be seen at two sites among some varieties.Phylogenetic tree constructed revealed the evolutionary relationships among these wheat varieties.[Conclusion] This study reveals the polymorphism and evolutionary characteristics in the nucleotide and amino acid sequences in different wheat varieties,which lays foundation for investigating the evolution and biological function of TaUBC4 gene.In addition,the phylogenetic tree constructed provides theoretical references for the classification of the wheat varieties with complicated background.

Identification and Comparative Analysis of DGAT1 Genes among Cotton and Other Model Plants

Diacylglycerol acyltransferase （DGAT） is the key enzyme that catalyzes the triacylglycerol biosynthesis. The comparative analysis was performed on the DGAT1 genes of cotton and other model plants. Sequence analysis showed that most of the DGAT1 genes,with high variation of intron length and high conservation of intron phrase,from the cotton and other model plants had 16 exons. Additionally, 7 conserved motifs were present in these DGAT1 proteins. The core motifs were overlapped with the functional domain of DGAT1 protein. Phylogenetic analysis demonstrated that gene tree was highly consistent to species tree,suggesting that the evolutionary history of species was revealed by gene tree. There was single copy of DGAT1 gene in cotton,but at least two duplicated DGAT1 genes were iden- tified in rice,maize,poplar and moss genomes. The selective pressure analysis showed that the PtDGATla/PtDGATlb was under positive selection,but other four pairs of homologous genes were under negative selection. 17 positively selected sites were identified at subgroup level （P〉0.05）,suggesting these subgroups under relaxed functional constraint. The findings provide a basis for further studying func- tion and evolution of DGAT1 genes in cotton and other model plants.

Evolution of COP9 Signalosome and Proteasome Lid Complex

The COP9 signalosome and the regulatory lid of the 26S proteasome are both eight-subunit protein complexes which are present in most eukaryotes. There is a one-to-one relationship between the corresponding subunits of the two protein complexes in terms of their size and amino acid sequences. Eight groups of subunits from the COP9 signalosome and the proteasome lid complex of different organisms are collected from all the databases at the NCBI website. The corresponding subunits of COP9 signalosome and proteasome lid complex share at least 12% amino acid identity and some conserved regions, and the conserved sites spread evenly over the entire length of the subunits, suggesting that the two complexes have a common evolutionary ancestor. Phylogenetic analyses based on the amino acid sequences of the corresponding subunits of two protein complexes indicate that every tree consists of two clades. The subunits from one of the two protein complexes of different organisms are grouped into one of the two clades respectively. The sequences of single-cell organisms are always the basal groups to that of multi-cell animal and plant species. These results imply that the duplication/divergence events of COP9 signalosome and regulatory lid of the proteasome genes have occurred before the divergence of single-cell and multi-cell eukaryotes, and the genes of the two complexes are independently evolved. The analyses of dN/dS correlation show significant Pearson's correlations between 21 and 15 pairs of subunit-encoding sequences within the COP9 signalosome and the proteasome lid complex respectively, suggesting that those subunits pairs might have related functions and interacted with one another, and resulted in co-evolution.

Cloning and Analysis of a Chalone Synthase Gene from Fagopyrum tataricum

[Objective] This study aimed to clone a full-length CHS gene from buck- wheat. [Method] With total RNA extracted from buckwheat as the template, CHS cD- NA sequence was cloned from buckwheat by using RACE technology and CODEHOP primer design method, the full length gene was obtained by primers which were de- signed for amplification of full-length gene sequence with buckwheat DNA template. Clustalxl.81 and MEGA4 software were used for sequence analysis and construction of phylogenetic tree; NCBI Blastn and Biastp programs were applied for homology analysis of nucleic acid and protein. [Result] Bioinformatics analysis showed that the full length of this gene is 1 906 bp, containing a 463 bp intron sequence and a 1 188 bp coding region, encoding 395 amino acids. Blastn sequence alignment revealed that the CHS gene sequence obtained in this study shared 86% homology with the CHS gene of closely related species. [Conclusion] This study laid the foundation to clarify molecular basis of the synthesis of buckwheat bioflavanoids and explore an effective way to improve the content of buckwheat bioflavanoids.

Cloning and Phylogenetic Analysis of a Fatty Acid Elongase Gene from Nannochloropsis oculata CS179

Nannochloropsis oculata CS179, a unicellular marine microalga, is rich in long-chain polyunsaturated fatty acids （LCPUFAs）. Elongase and desaturase play a key role in the biosynthesis of PUFAs. A new elongase gene, which encodes 322 amino acids, was identified via RT-PCR and 5＇ and 3＇ RACE. The sequence of the elongase gene was blast-searched in the NCBI GenBank and showed a similarity to those of the coptosporidium. But the NJ-tree revealed that the N. oculata CS 179 elongase clustered with those of the microalgae Phaeodac^lum tricornutum, Ostreocoecus tauri and Thalassiosira pseudonana.

Effect of Using Baker＇s Yeast and Exogenous Digestive Enzymes as Growth Promoters on Growth, Feed Utilization and Hematological Indices of Nile tilapia, Oreochromis niloticus Fingerlings

The present study was conducted to evaluate the effect of baker yeast, Saccharomyces cerevisiae （SC） and exogenous digestive enzymes （pepsin, papain and a-amylase, EDE） dietary supplementation on growth performance, feed utilization and hematological indices of Nile tilapia, Oreochromis niloticus fingerlings. A total of 630 Nile tilapia fingerlings with an average body weight of 26.4 ± 0.2 g were divided in the seven experimental net-pen treatments （three replicates each）. The experiment was conducted for 119 days. Seven isonitrogenous （26.50%） digestible protein and isocaloric （13.40 MJ kgl） digestible energy experimental diets were formulated. The control diet had no SC and EDE added. Diets 2-3 each contained SC at levels of 2 and 4 g 100 g diet-t, respectively, while diets 4-5 each contained EDE at levels of （0.64, 1.28, 0.16） and （1.28, 2.56, 0.32） g 100 gdiefI of pepsin, papain and a-amylase, respectively. Diet 6 contained mixture of SC and EDE at levels of 1 g yeast and 0.32, 0.64, 0.08 g of pepsin, papain and a-amylase, respectively 100 gdiet1 and diet D7 contained 2 g yeast and 0.64, 1.28, 0.16 g of pepsin, papain and a-amylase, respectively 100 g dietl. Growth performance and feed utilization efficiency of Nile tilapia were significantly （P 〈 0.05） higher in all treatments receiving SC and/or EDE supplemented-diets than the control diet which suggests that the addition of SC and EDE enhanced the growth performance. Red blood cells counts, hematocrit and hemoglobin were significantly （P 〈 0.05） highest in all treatments receiving mixture of SC and EDE supplemented-diets （D6 ＋ D7）. The same trend was observed for total plasma protein and total plasma globulin levels. The results of present study suggested that Nile tilapia fingerlings fed diets containing the mixture of I g yeast, SC and 0.32, 0.64, 0.08 g of pepsin, papain and a-amylase, respectively 100 gdiet^-1, for 119 days had enhanced growth performance, diet utilization efficiency and hematological indices.

Directed Molecular Evolution of Nitrite Oxido-reductase by DNA-shuffling

Objective To devellop directly molecular evolution Of nitrite oxido-reductase using DNA-shuffling technique because nitrobacteria grow extremelly slow and are unable to nitrify effectively inorganic nitrogen in wastewater treatmem. Methods The norB gene coding the ntitrite oxido-reductase in nitrobacteria was cloned and sequenced. Then, directed molecular evolution of nitrite oxido-reductase was developed by DNA-shuffling of 15 norB genes from different nitrobacteria. Results After DNA-shuffling with sexual PeR and staggered extension process PCR, the sequence was differem from its parental DNA fragmems and the homology ranged from 98% to 99%. The maximum nitrification rate of the modified bacterium of X16 by DNA-shuffling was up to 42.9 mg/L.d, which was almost 10 times higher than that of its parental bacteria. Furthermore, the modified bacterium had the same characteristics of its parental bacteria of E. coli and could grow rapidly in normal cultures. Conclusion DNA-shuffling was successfully used to engineer E. coli, which had norB gene and could degrade inorganic nitrogen effectively.

Green Synthesis and Physical and Chemical Characterization of Chitosans with a High Degree of Deacetylation, Produced by a Binary Enzyme System

Chitosan is a biopolymer obtained from chitin, where the N-acetylglucosamine monomer is in its deacetylated form; this polymer is useful for a wide variety of industrial applications. The properties and uses of chitosan depend on its physical and chemical characteristics, which result from the treatments used for its production. In this study, we report the preparation and characterization ofchitosan oligosaccharides by a green synthesis from crystalline shrimp chitin, using a sequential enzyme treatment by chitinase and chitin deacetylase. Chitinases were purified from grapes and used to rupture the crystalline shrimp chitin structure, modifying the crystallinity index from 57.6% to 15.9%. The resultant polymers were deacetylated using a recombinant chitin deacetylase from Saccharomyces cerevisiae, which was cloned and expressed in Pichia pastoris. The chitosans produced showed an estimated DA （degree of acetylation） of approximately 20%, and the molecular weights ranged from -7,600 to -3,700 after treatment in pH 3.0 and pH 6.0 for 10 min and 40 min, respectively. Physical and chemical characterization of the products indicated that enzyme fragmentation of chitin probably makes the acetamide groups more accessible to deacetylation, forming homogeneous polymers that are free of hazardous sub-products, have defined low molecular weights, and are highly deacetylated.

Mitochondrial pyruvate dehydrogenase E1 of Nosema bombycis:A marker in Microsporidian evolution

Microsporidia are a group of intracelluar eukaryotic parasites, which can infected almost all animals, including human beings. Till now, no mitochodria but mitosome, a remnant of mitochondria was discovered in this phylum. We present here the mitochondrial pyruvate dehydrogenase El （PDH, including PDHα and PDHβ） of the microsporidian Nosema bombycis, the pathogen of silkworm pebrine. Compared with PDH of microsporidian Encephalitozoon cuniculi and Antonospora locustae, both subunits are eonscrced. The phylogeny indicated that both subunits are mitochondrial. The syntenic maps revealed the subunits organization of NbPDH is distributed in different scaffolds, similar to that of EcPDH but different with AIPDH, and the relationship between phylogeny tree and organization of PDH suggest that the AlPDH subunits organization is the ancestral style of microsporidia, and through the genome evolution, the reshuffling of the chromosome of microsporidia occurred, the adjacent style of ALPDHE1 organization changed, and the two subunits separated and located to different chromosomes in E. cuniculi. For N. bombycis and N. ceranae, they locate to different scaffolds. In order to determine NbPDH subcellular localizations, we prepared the polyclonal antibodies against NbPDH prokaryotic fusion proteins, and adopted the colloidal gold immunological electron microscopy, the expression signals of NbPDH were observed in spores however, the subcellular localization were not definited. In general, through comparison of three mierosporidian PDH molecular phylogeny, subunits organization in chromosomes, localization indicated that PDH is an interesting marker in microsporidia evolution

Xanthophyll Cycle and Its Relative Enzymes

Light is a fundamental source of energy but is also potentially harmful to organisms. Plants have evolved a variety of regulatory mechanisms to respond to the naturally varying light conditions. Xanthophyll cycle is now recognized as a key regulator and photoprotective mechanism found ubiquitously in plants. Xanthophyll cycle has multiple functions, such as thermal dissipation, protection against oxidative stress caused by light, modulation of the structure of thylakoid membrane, involving in blue light signal transduction and regulating the synthesis ofABA （Abscisic acid）. VDE （Violaxanth de-epoxidase） and ZE （zeaxanth epoxidase）, are involved in xanthophyll cycle. This paper outlined the functions of xanthophylls cycle and its relative enzymes.

Microevolution of Mitochondrial Cytochrome oxidase subunit I in Drosophila melanogaster at ＂Evolution Canyon＂, Israel

We determined the sequence of mitochondrial Cytochrome oxidase subunit I （CO1） in two Drosophila melanogaster strains originating at ＂Evolution Canyon＂, Israel. CO1 nucleotide sequences from two iso-female strains, 2-1 and 6-1, were 1,534 and 1,543 base-pairs, respectively. In each strain, ATAA was used in initiation of translation. Exchange rates for nucleotide and amino acid sequences were larger in the 6-1 strain than in the 2-1 strain when Oregon-R was used as the standard. Non-synonymous exchange rate was larger than synonymous exchange rate among the three strains.

Detection of plasmid-mediated IMP-1 metallo-β-lactamase and quinolone resistance determinants in an ertapenem-resistant Enterobacter cloacae isolate

Objective： To investigate the mechanism of carbapenem resistance and the occurrence ofplasmid-mediated quinolone resistance determinants qnr and aac（6＇）-Ib-cr in a clinical isolate of Enterobacter cloacae. Methods： An ertapenem-resistant E. cloacae ZY106, which was isolated from liquor puris of a female gastric cancer patient in a Chinese hospital, was investigated. Antibiotic susceptibilities were determined by agar dilution method. Conjugation experiments, isoelectric focusing, polymerase chain reaction （PCR）, and DNA sequence analyses of plasmid-mediated carbapenemases and quinolone resistance determinants were preformed to confirm the genotype. Outer membrane proteins （OMPs） were examined by urea-sodium dodecyl sulfate- polyacrylamide gel electrophoresis （Urea-SDS-PAGE）. Results： Minimum inhibitory concentrations （MICs） of imipenem, mer- openem, and ertapenem for ZY 106 were 2, 4, and 16 pg/ml, respectively. Conjugation studies with Escherichia coli resulted in the transfer of significantly reduced carbapenem susceptibility. ZY106 produced IMP-1 metallo-β-lactamase and CTX-M-3 extended-spectrum β-1actamase, and E. coli transconjugant produced IMP-1. Plasrnid-mediated quinolone resistance determinant qnrS1 was detected in ZY106. Transfer of the qnrSl-encoding-plasmid into E. coli by conjugation resulted in intermediate resistance to ciprofloxacin in E. coli transconjugant. Urea-SDS-PAGE analysis of OMPs showed that ZYI06 lacked an OMP of approximately 38 kDa. Conclusion： It is the first IMP-l-producing Enterobacteriaceae in China and the first report of a clinical isolate that harbors both blalMP and qnrS genes as well. The blalMP-1, blaCTX-M-3, and qnrS1 are encoded at three different plasmids. IMP-1 combined with the loss of an OMP possibly resulted in ertapenem resistance and reduced imipenem and mero- penern susceptibility in E. cloacae.

Route improvement of 3-substituted-4-(2-methylcyclohexyloxy)-6-phenethylpyridinone

trans-3-Isopropyl-4-（2-methylcyclohexyloxy）-6-phenethylpyridin-2（1H）-one, as reverse transcriptase （NNRTIs）, exhibited significant potent activity not only against wild-type HIV-1 strains but also on mutant strains. For furthering study this compound, the original synthetic route should be shorten to improve the total yield. In this report, we designed an efficient synthetic strategy to obtain the target compound with higher yield.