Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase c DNA of Paphia textile（Pt CAT） was clo...Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase c DNA of Paphia textile（Pt CAT） was cloned using RTPCR and rapid amplification of c DNA ends（RACE）. Pt CAT is 1 921 bp long and consists of a 5′-UTR of 50 bp, a 3′-UTR of 349 bp, and an ORF of 1 542 bp that encodes 513 amino acids with a molecular weight of 58.4 k D and an estimated isoelectric point of 8.2. Sequence alignment indicated that Pt CAT contained a highly conserved catalytic signature motif（^（61）FNRERIPERVVHAKGAG^（77））, a proximal heme-ligand signature sequence（^（352）RLFSYSDP^（359））, and three catalytic amino acid residues（H^（72）, N^（145）, and Y^（356））. Pt CAT also contains two putative N-glycosylation sites（^（34）NKT^（36） and ^（437）NFT^（439）） and a peroxisome-targeting signal（^（511）AQL^（513））. Furthermore, Pt CAT shares 53%–88% identity and 29%–89% similarity with other catalase amino acid sequences. Pt CAT m RNA was present in all tested organs, including the heart, digestive gland, adductor muscle, gonad, gill, and mantle, but its expression was highest in the digestive gland. High-temperature-induced stress produced two expression patterns of Pt CAT m RNA： first, an initial up-regulation followed by a down-regulation in the heart, digestive gland, and gonad and, second, consistent down-regulation in all other organs. These results demonstrate that Pt CAT is a typical member of the catalase family and might be involved in the responses to harmful environmental factors.展开更多
Water is a major limiting factor for food production and many countries fail to produce sufficient food for their population due to severe water scarcity （Jury and Vaux, 2005）. Rice is the main staple food worldwide...Water is a major limiting factor for food production and many countries fail to produce sufficient food for their population due to severe water scarcity （Jury and Vaux, 2005）. Rice is the main staple food worldwide. More than 50% of rice in the world is rain-fed and drought causes severe reduction in rice grain yield in rain-fed environments （Venuprasad et al., 2007; Zhang, 2007; Sandhu et al., 2014）. Therefore, enhancing drought resistance （DR） of rice is important for food security. However, DR is a complex trait, which is controlled by a large number of loci with small effect and is also affected by different genetic background, genotype-by-environment interaction and other stresses such as heat （Hu and Xiong, 2014）.展开更多
The full-length Mlo gene was obtained by reverse transcription polymerase chain reaction （RT-PCR） and RACE. The result of sequence analysis indicated that M/o gene from Pericallis hybrida B. Nord. contained about 12...The full-length Mlo gene was obtained by reverse transcription polymerase chain reaction （RT-PCR） and RACE. The result of sequence analysis indicated that M/o gene from Pericallis hybrida B. Nord. contained about 1296bp open reading frame and encoded 431 amino acids. According to the comparison of the exogenous gene sequences by BLAST analysis and phylogenetic analysis, Mlo gene shared over 85% nucleotide homology and 98% amino acid homology. Finally, through semi-quantitative-PCR and fluorescence quantitative analysis, we found that Mlo gene showed the highest expression levels in leaves and the lowest in roots after inoculated with powdery mildew pathogen for different days.展开更多
ESTs fragments which represents corresponding novel genes were obtained by sequencing and bioinformatics analysis of human fet al kidney cDNA library. Microarray was prepared by using these novel EST fragmen ts by a...ESTs fragments which represents corresponding novel genes were obtained by sequencing and bioinformatics analysis of human fet al kidney cDNA library. Microarray was prepared by using these novel EST fragmen ts by automatic spotting. Expression patters of 79 ESTs of novel genes from huma n fetal kidney were analyzed in fetal brain and fetal heart tissues of 20\|week\ | and 26\|week\|age fetus by performing of cDNA chip hybridization. This provide s clues for studying exact functions of the novel genes. 8 genes were obtained w hich were expressed differentially in the fetal brain and heart of 20\|week\| an d 26\|week\|age respectively. Then differentially expressed genes were identifie d by Northern analysis. The more exact function of the novel genes is under stud y.展开更多
In order to confirm the existence of indoleamine 2,3-dioxygenase(IDO) gene in swine,and to clone the novel gene followed by the molecule structure properties and expression pattern analysis,the porcine mRNA sequences ...In order to confirm the existence of indoleamine 2,3-dioxygenase(IDO) gene in swine,and to clone the novel gene followed by the molecule structure properties and expression pattern analysis,the porcine mRNA sequences homologous to human IDO were obtained from GenBank database by bioinformatics method.By using RT-PCR,the IDO gene was cloned from porcine endothelial cell line and the accuracy of the nucleic acid sequence was confirmed,and the expression pattern of the gene was detected.The three-dimensional structure model of porcine IDO was built referring to the tertiary structure of human IDO using biological sequence analysis software and database.The results showed that the porcine IDO was identified by sequencing.The nucleotide sequences were confirmed as a novel gene after submitted to Genbank.Porcine IDO was expressed in the lung,thymus,epididymis and anterior chamber with a basic level,however in peripheral blood mononuclear cells(PBMCs) the IDO gene was highly expressed.The three-dimensional structure model of porcine IDO was similar to that of human IDO.It was suggested that identification of the structure information of porcine IDO is essential to further investigate the immunologic function of the gene.Study of IDO on NK cells-mediated xenograft rejection will be a novel therapeutic target for the development of xenotransplantation.展开更多
Objective: To identify the differential expression profile of human novel gene UBAP1, a putative nasopharyngeal neoplasms (NPC) relate gene, in multiple cancers. Methods: We first present an EST approach for electroni...Objective: To identify the differential expression profile of human novel gene UBAP1, a putative nasopharyngeal neoplasms (NPC) relate gene, in multiple cancers. Methods: We first present an EST approach for electronic Northern in silico to analyse expression patterns of UBAP1 in tumor and normal tissues. Full length cDNA of UBAP1 gene was taken as a “probe” sequence, and a blastn search was performed against human EST Database. The Blastn report can be used to determine the fold differences between the pedigree ESTs in different libraries. Especially, the ESTs corresponding to UBAP1 present in fifteen tumor-derived libraries were compared against their normal counterpart to produce an electronic differential expression profile. Second, the distinct down-regulation of UBAP1 in meningioma, glioma, and colorectal tumor was confirmed by differentially RT-PCR analysis. Results: Database surveys indicated that UBAP1 gene was not only ubiquitously expressed in many normal tissues with various levels but also differentially expressed in different tumor tissues, especially down-regulated in multiple neoplastic tissues such as brain, breast, skin, colon, testis and uterus tumor tissues. Furthermore, differential RT-PCR analysis demonstrated that expression of UBAP1 was down-regulated or absent in 7 of 12 (58%) meningioma samples, 6 of 9 (66%) glioma and 7 of 11 (63%) colorectal tumor tissues respectively. Conclusion: we described a data mining procedure in silico that proved to be useful for the identification of differential expression patterns of UBAP1. These findings could be valuable for the investigation of the mechanism the differential expression of UBAP1 gene and its significance in the progression of multiple cancers.展开更多
A repeated sequence DNA fragment, L5B-4, was cloned from the 5 kb BamHI DNA fragments of rat genomic DNA. The expressions of the L5B-4 DNA fragment are different in liver and hepatoma cells. The amounts of transcripts...A repeated sequence DNA fragment, L5B-4, was cloned from the 5 kb BamHI DNA fragments of rat genomic DNA. The expressions of the L5B-4 DNA fragment are different in liver and hepatoma cells. The amounts of transcripts in hepatoma cells are lower in nucleus and higher in cytoplasm, especially in polysomal RNA, as compared with that in liver cells. The alteration shown in polysomal RNA of hepatoma cells seems to be specific. These results are discussed with respect to the possible function of this repeated DNA and its variation in hepatoma cells.展开更多
Rhododendron simsii（Ericaeae：Rhododendron） has high ornamental value and ecological value.In this study,7 pairs of novel EST-SSR markers were developed from the genomic sequence of R.simsii,and they were used to in...Rhododendron simsii（Ericaeae：Rhododendron） has high ornamental value and ecological value.In this study,7 pairs of novel EST-SSR markers were developed from the genomic sequence of R.simsii,and they were used to investigate the genetic diversity of 32 natural R.simsii samples from Guifeng Mountain,Hubei Province.Results showed that a total of 31 polymorphic bands were amplified with allele number per locus of 4.43.Mean values of heterozygosity（Ho） and expected heterozygosity（He） were 0.679 58 and 0.723 14,respectively.This research will not only enrich the existing SSR database,but also lay a foundation for subsequent studies about molecular marker-assisted breeding,genetic diversity analysis,genetic structure analysis and phylogenetic analysis.展开更多
Powdery mildew is a serious disease of wheat in China. As part of ITEC (International Triteace EST cooperation), EST (expressed sequence tags) technique was used to explore the gene expression in leaf induced by Ery...Powdery mildew is a serious disease of wheat in China. As part of ITEC (International Triteace EST cooperation), EST (expressed sequence tags) technique was used to explore the gene expression in leaf induced by Erysiphe graminis DC. A conventional cDNA library was constructed, and a total of 1 500 clones picked randomly from the library were sequenced, three hundred and eighty_seven ESTs of them were unique, which got the Accession Number in GenBank. About 49.4% ESTs showed significant similarity to functions of known sequences in GenBank. There are 196 ESTs' with functions not able to be determined, and eighty_four ESTs were demonstrated to be novel sequences. High_density dot membranes from unique clones were produced, and several disease resistance related genes were screened by differential hybridization.展开更多
Objective: To identify a cDNA clone from the subtracted library of human hepatocellular carcinoma (HCC). Methods: Suppression subtractive hybridization was used to isolated a panel of genes that are differentially exp...Objective: To identify a cDNA clone from the subtracted library of human hepatocellular carcinoma (HCC). Methods: Suppression subtractive hybridization was used to isolated a panel of genes that are differentially expressed in hepatocellular carcinoma as compared with cirrhotic liver. T/A cloning method was used to construct a subtracted cDNA library. DNA sequencing analysis and Northern blot analysis were also utilized. Results: The cloned cDNA is 787 nucleotides in length and contains an open reading frame of 230 amino acids, which is a cDNA fragment of reported human fibrinogen, gamma polypeptide (FGG). Northern analysis revealed that this gene was overexpressed in two hepatocellular carcinoma cell lines, SMMC-7721 and HepG2. Conclusion: Sequence identity proved the cDNA clone fragment of as FGG gene. Differential expression of the cDNA fragment in HCC suggested that FGG is related to HCC, indicating a new clue for developing a novel diagnostic and prognostic marker.展开更多
Seventy-five previously known plant microRNAs (miRNAs) were classified into 14 families according to their gene sequence identity. A total of 18,694 plant expressed sequence tags (EST) were found in the GenBank EST da...Seventy-five previously known plant microRNAs (miRNAs) were classified into 14 families according to their gene sequence identity. A total of 18,694 plant expressed sequence tags (EST) were found in the GenBank EST databases by comparing all previously known Arabidopsis miRNAs to GenBank’s plant EST databases with BLAST algorithms. After removing the EST sequences with high numbers (more than 2) of mismatched nucleotides, a total of 812 EST contigs were identified. After predicting and scoring the RNA secondary structure of the 812 EST sequences using mFold software, 338 new potential miRNAs were identified in 60 plant species. miRNAs are widespread. Some microRNAs may highly conserve in the plant kingdom, and they may have the same ancestor in very early evolution. There is no nucleotide substitution in most miRNAs among many plant species. Some of the new identified potential miRNAs may be induced and regulated by environmental biotic and abiotic stresses. Some may be preferentially expressed in specific tissues, and are regulated by developmental switching. These findings suggest that EST analysis is a good alternative strategy for identifying new miRNA candidates, their targets, and other genes. A large number of miRNAs exist in different plant species and play important roles in plant developmental switching and plant responses to environmental abiotic and biotic stresses as well as signal transduction. Environmental stresses and developmental switching may be the signals for synthesis and regulation of miRNAs in plants. A model for miRNA induction and expression, and gene regulation by miRNA is hypothesized.展开更多
The development of expressed sequence tag-derived simple sequence repeats(EST-SSRs) provided a useful tool for investigating plant genetic diversity.In the present study,22 polymorphic EST-SSRs from grain soybean were...The development of expressed sequence tag-derived simple sequence repeats(EST-SSRs) provided a useful tool for investigating plant genetic diversity.In the present study,22 polymorphic EST-SSRs from grain soybean were identified and used to assess the genetic diversity in 48 vegetable soybean accessions.Among the 22 EST-SSR loci,tri-nucleotides were the most abundant repeats,accounting for 50.00% of the total motifs.GAA was the most common motif among tri-nucleotide repeats,with a frequency of 18.18%.Polymorphic analysis identified a total of 71 alleles,with an average of 3.23 per locus.The polymorphism information content(PIC) values ranged from 0.144 to 0.630,with a mean of 0.386.Observed heterozygosity(H o) values varied from 0.0196 to 1.0000,with an average of 0.6092,while the expected heterozygosity(H e) values ranged from 0.1502 to 0.6840,with a mean value of 0.4616.Principal coordinate analysis and phylogenetic tree analysis indicated that the accessions could be assigned to different groups based to a large extent on their geographic distribution,and most accessions from China were clustered into the same groups.These results suggest that Chinese vegetable soybean accessions have a narrow genetic base.The results of this study indicate that EST-SSRs from grain soybean have high transferability to vegetable soybean,and that these new markers would be helpful in taxonomy,molecular breeding,and comparative mapping studies of vegetable soybean in the future.展开更多
Zostera marina, a monocotyledonous angiosperm, is one of the most important seagrass species. To inves- tigate the salt-tolerance mechanism and discover salt-tolerant genes in Z. marina, a cDNA library was con- struct...Zostera marina, a monocotyledonous angiosperm, is one of the most important seagrass species. To inves- tigate the salt-tolerance mechanism and discover salt-tolerant genes in Z. marina, a cDNA library was con- structed. Single-pass sequencing of the 5＇ ends of 4 081 clones yielded 4 002 high quality expressed sequence tags （ESTs）, which were assembled into 241 contigs and 1 673 singletons, representing 1 914 unigenes. The average length of the ESTs was 582 bp, with sizes ranging from 100-1 500 bp. Basic Local Alignment Search Tool （BLASTX） analysis revealed that 1 664 unigenes had significant homology to known genes in the Na- tional Center for Biotechnology Information （NCBI） non-redundant （nr） database （E-value≤5-10）. Among them, the two most abundant genes encoded metallothionein （157 ESTs） and chlorophyll a/b-binding pro- tein （38 ESTs）, accounting for 7.1% and 1.7% of the total ESTs, respectively. Using Kyoto Encyclopedia of Genes and Genomes （KEGG）, 1 462 unigenes were assigned to 1 161 pathways （E-value≤5-10）. A total of 938 unigenes were assigned Gene Ontology （GO） terms based on the GO hierarchy analysis, and InterProScan searches recognized 1 003 InterPro families. Three genes for metallothionein in Z. marina that belonged to Class II was identified. Results of this study will improve understanding of the molecular mechanisms of saline tolerance in Z. marina.展开更多
Paris polyphylla Smith var.yunnanensis(Franch.) Hand.-Mazz.is a rhizomatous,herbaceous,perennial plant that has been used for more than a thousand years in traditional Chinese medicine.It is facing extinction due to o...Paris polyphylla Smith var.yunnanensis(Franch.) Hand.-Mazz.is a rhizomatous,herbaceous,perennial plant that has been used for more than a thousand years in traditional Chinese medicine.It is facing extinction due to overharvesting.Steroids are the major therapeutic components in Paris roots,the commercial value of which increases with age.To date,no genomic data on the species have been available.In this study,transcriptome analysis of an 8-year-old root and a 4-year-old root provided insight into the metabolic pathways that generate the steroids.Using Illumina sequencing technology,we generated a high-quality sequence and demonstrated de novo assembly and annotation of genes in the absence of prior genome information.Approximately 87,577 unique sequences,with an average length of 614 bases,were obtained from the root cells.Using bioinformatics methods,we annotated approximately 65.51% of the unique sequences by conducting a similarity search with known genes in the National Center for Biotechnology Information's non-redundant database.The unique transcripts were functionally classified using the Gene Ontology hierarchy and the Kyoto Encyclopedia of Genes and Genomes database.Of 3082 genes that were identified as significantly differentially expressed between roots of different ages,1518(49.25%) were upregulated and 1564(50.75%) were downregulated in the older root.Metabolic pathway analysis predicted that 25 unigenes were responsible for the biosynthesis of the saponins steroids.These data represent a valuable resource for future genomic studies on this endangered species and will be valuable for efforts to genetically engineer P.polyphylla and facilitate saponin-rich plant development.展开更多
Objective： To screen and analyze key express sequence tags （ESTs） which were differentially displayed in every period of SD rats＇ primary hepatic carcinoma and reveal the molecular mechanism of carcinogenesis. Met...Objective： To screen and analyze key express sequence tags （ESTs） which were differentially displayed in every period of SD rats＇ primary hepatic carcinoma and reveal the molecular mechanism of carcinogenesis. Methods： Using diethylnitrosamine （DENA） as a cancerigenic agent, animal models with different phases of primary hepatic cancer were constructed in SD rats. Rats were respectively sacrificed at d 14, d 28, d 56, d 77, d 105 and d 112 after the rats received DENA by gavage, then the livers were harvested. One part of the livers was classified according to their pathological changes, while the other was reserved for molecular mechanism studies on hepatocarcinogenesis. The differentially expressed genes were isolated from both normal and morbid tissues by mRNA differential display technique （DDRT-PCR）. After the fragments were sequenced, bioinformatics were .used to analyze the results. Results： Twelve differentially expressed cDNA fragments were obtained. Nine fragments had the homology with known cDNA clones, especially EST-7 was similar to BN/SsNHsdMCW mitochondrion gene and the identity was 100% which suggested EST-7 may be the part of BN/SsNHsdMCW mitochondrion gene. In contrast, other three fragments （EST-1, EST-3 and EST-5） had extremely low identity to any genes registered in GENBANK databases. Conclusions： BN/SsNHsdMCW mitochondrion gene was expressed in different periods of hepatocarcinogenesis. Moreover, EST-I, EST-3 and EST-5 were suggested to contribute to the development of rat hepatocarcinogenesis, and thus may be candidates of new targets of oncogenes or cancer suppressor genes.展开更多
Faba bean （Vicia faba L.）, one of the most important legumes in the world, evolved different types of cultivars due to its partial cross-pollination. The development of simple sequence repeat （SSR） markers from ex...Faba bean （Vicia faba L.）, one of the most important legumes in the world, evolved different types of cultivars due to its partial cross-pollination. The development of simple sequence repeat （SSR） markers from expressed sequence tags （EST） provided a useful tool for investigation of its genetic diversity. The purpose of the present study was to investigate the genetic diversity of faba bean from China and Europe using EST-SSR markers. 5 031 faba bean ESTs from the NCBI database were downloaded and assembled into 1 148 unigenes. A total of 107 microsatellites in 96 unigenes were identified, indicating that merely 8.36% of sequences contained SSRs. The most abundant SSR within faba bean was tri-nucleotide repeat motif, and among all the tri-nucleotide repeats, the motif AAG/CTT was the most abundant type. Based on these results, 11 EST-SSR markers were used to assess the genetic diversity of 29 faba bean cultivars from China and Europe with two to three alleles per locus. The polymorphism information content value ranged from 0.0644 to 0.4278 with an average of 0.2919. Principal coordinate analysis （PCA） and phylogenetic clustering based on these 11 EST-SSR markers distinguished these cultivars into different groups. The results indicated that faba bean in China had a narrow genetic basis, and the additional sources of genetic cultivars/accessions should be introduced to enhance the genetic variability. The results of this study proved that the EST-SSR marker is very effective in evaluation of faba bean germplasm.展开更多
In 2008, a green tide broke out before the sailing competition of the 29th Olympic Games in Qingdao. The causative species was determined to be Enteromorpha prolifera （Ulva prolifera O. F. Miiller）, a familiar green...In 2008, a green tide broke out before the sailing competition of the 29th Olympic Games in Qingdao. The causative species was determined to be Enteromorpha prolifera （Ulva prolifera O. F. Miiller）, a familiar green macroalga along the coastline of China. Rapid accumulation of a large biomass of floating U. prolifera prompted research on different aspects of this species. In this study, we constructed a nonnormalized cDNA library from the thalli of U. prolifera and acquired 10072 high-quality expressed sequence tags （ESTs）. These ESTs were assembled into 3 519 nonredundant gene groups, including 1 446 clusters and 2 073 singletons. After annotation with the nr database, a large number of genes were found to be related with chloroplast and ribosomal protein, GO functional classification showed 1 418 ESTs participated in photosynthesis and 1 359 ESTs were responsible for the generation of precursor metabolites and energy. In addition, rather comprehensive carbon fixation pathways were found in U. prolifera using KEGG. Some stress-related and signal transduction-related genes were also found in this study. All the evidences displayed that U. prolifera had substance and energy foundation for the intense photosynthesis and the rapid proliferation. Phylogenetic analysis of cytochrome c oxidase subunit I revealed that this green-tide causative species is most closely affiliated to Pseudendoclonium akinetum （Ulvophyceae）.展开更多
Aluminum (Al) toxicity is the major factor limiting crop productivity in acid soils. In this study, a recombinant inbreed line (RIL) population derived from a cross between an Al sensitive lowland indica rice variety...Aluminum (Al) toxicity is the major factor limiting crop productivity in acid soils. In this study, a recombinant inbreed line (RIL) population derived from a cross between an Al sensitive lowland indica rice variety IR1552 and an Al tolerant upland japonica rice variety Azucena, was used for mapping quantitative trait loci (QTLs) for Al tolerance. Three QTLs for relative root length (RRL) were detected on chromosome 1, 9, 12, respectively, and 1 QTL for root length under Al stress is identical on chromosome 1 after one week and two weeks stress. Comparison of QTLs on chromosome 1 from different studies indicated an identical interval between C86 and RZ801 with gene(s) for Al tolerance. This interval provides an important start point for isolating genes responsible for Al tolerance and understanding the genetic nature of Al tolerance in rice. Four Al induced ESTs located in this interval were screened by reverse Northern analysis and confirmed by Northern analysis. They would be candidate genes for the QTL.展开更多
MicroRNAs(miRNA)are recently discovered endogenous,small noncoding RNAs(of 22 nucleotides)that play pivotal roles in gene regulation.They are involved in post-transcriptional control of gene expression.miRNAs are emer...MicroRNAs(miRNA)are recently discovered endogenous,small noncoding RNAs(of 22 nucleotides)that play pivotal roles in gene regulation.They are involved in post-transcriptional control of gene expression.miRNAs are emerging as important regulators of cell proliferation,development,cancer formation,stress responses,cell death and physiological conditions.Increasing evidence has demonstrated the human miRNAs bind to their target mRNA sequences with perfect or near-perfect sequence complementarily.This provides a powerful strategy for discovering potential type 2 diabetes mellitus(T2DM)targets and gives the probability to exploit them for diagnostic and therapeutic causes.About 6%of the world population is affected by T2DM,and it is recognized as a global epidemic by the World Health Organization.At present there is no valid biomarker to control or manage T2DM.Therefore,the present study applied a mature sequence of miRNAs from publicly accessible databases to identify the miRNA from T2DM expressed sequence tags,and the results are detailed and discussed below.展开更多
SSR markers are useful in pepper linkage mapping and gene location.446 SSR markers have been reported,but they are insufficient.It is costly to develop SSR markers from DNA library,whereas it seems much easy to find i...SSR markers are useful in pepper linkage mapping and gene location.446 SSR markers have been reported,but they are insufficient.It is costly to develop SSR markers from DNA library,whereas it seems much easy to find in EST sequences in the GenBank of pepper through internet.In this study,attempts have been made to develop SSR markers in the EST sequences by using bioinformatics.EST sequences were trimmed by‘est-trimmer.pl’software,while 116915 EST sequences were obtained without poly‘A’or poly‘T’,ranged between 100 and 700 bp.Using‘e-PCR’and‘del.pl’softwares,SSR sequences were identified.2508 microsatellite loci(larger than 20 repeats)were established and 755 SSR primers were designed using SSR finder software and Primer 3 software.There were 498(0.43%)mono-,1026(0.89%)di-,518(0.45%)tri-,245(0.21%)tetra-,114(0.10%)penta-,and 107(0.09%)hexa-nucleotide SSRs.The estimated frequency of SSRs was approximately 1/25.12 kb.According to the distribution of SSRs in pepper,the mean length of pepper SSRs was 22.68 bp and the adenine rich repeats such as A/T,AG,AT,AAG,AAAT,and AAAC were predominant in each type of SSRs(mono-,di-,tri-,tetra-,penta-,and hexa-),whereas the C/G,CG,CCG repeats were less abundant.210 primers were tested in 8 pepper cultivars and the PCR result revealed the existence of polymorphism among 127(60.48%)SSR primers within 8 pepper cultivars.It is confirmed that pepper EST database could be efficiently exploited for availability of SSR markers.展开更多
基金The National Natural Science Foundation of China under contract No.31172397the New Century Excellent Talents of Fujian Province University under contract No.JA14167the Open Research Fund Program of Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment under contract No.Z814041
文摘Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase c DNA of Paphia textile（Pt CAT） was cloned using RTPCR and rapid amplification of c DNA ends（RACE）. Pt CAT is 1 921 bp long and consists of a 5′-UTR of 50 bp, a 3′-UTR of 349 bp, and an ORF of 1 542 bp that encodes 513 amino acids with a molecular weight of 58.4 k D and an estimated isoelectric point of 8.2. Sequence alignment indicated that Pt CAT contained a highly conserved catalytic signature motif（^（61）FNRERIPERVVHAKGAG^（77））, a proximal heme-ligand signature sequence（^（352）RLFSYSDP^（359））, and three catalytic amino acid residues（H^（72）, N^（145）, and Y^（356））. Pt CAT also contains two putative N-glycosylation sites（^（34）NKT^（36） and ^（437）NFT^（439）） and a peroxisome-targeting signal（^（511）AQL^（513））. Furthermore, Pt CAT shares 53%–88% identity and 29%–89% similarity with other catalase amino acid sequences. Pt CAT m RNA was present in all tested organs, including the heart, digestive gland, adductor muscle, gonad, gill, and mantle, but its expression was highest in the digestive gland. High-temperature-induced stress produced two expression patterns of Pt CAT m RNA： first, an initial up-regulation followed by a down-regulation in the heart, digestive gland, and gonad and, second, consistent down-regulation in all other organs. These results demonstrate that Pt CAT is a typical member of the catalase family and might be involved in the responses to harmful environmental factors.
基金supported by grants from the National Program for Basic Research of China(No.2012CB114305)the National Program on High Technology Development(No. 2012AA10A303)the Oversea Graduate Program from Ministry of Education to K.Songyikhangsuthor
文摘Water is a major limiting factor for food production and many countries fail to produce sufficient food for their population due to severe water scarcity （Jury and Vaux, 2005）. Rice is the main staple food worldwide. More than 50% of rice in the world is rain-fed and drought causes severe reduction in rice grain yield in rain-fed environments （Venuprasad et al., 2007; Zhang, 2007; Sandhu et al., 2014）. Therefore, enhancing drought resistance （DR） of rice is important for food security. However, DR is a complex trait, which is controlled by a large number of loci with small effect and is also affected by different genetic background, genotype-by-environment interaction and other stresses such as heat （Hu and Xiong, 2014）.
基金Supported by Postdoctoral Scientifi c Research Foundation of Heilongjiang Province(LBH-Q10144)the Natural Science Foundation of Heilongjiang Province(C201112)Northeast Agricultural University Doctoral Research Fund(200830)
文摘The full-length Mlo gene was obtained by reverse transcription polymerase chain reaction （RT-PCR） and RACE. The result of sequence analysis indicated that M/o gene from Pericallis hybrida B. Nord. contained about 1296bp open reading frame and encoded 431 amino acids. According to the comparison of the exogenous gene sequences by BLAST analysis and phylogenetic analysis, Mlo gene shared over 85% nucleotide homology and 98% amino acid homology. Finally, through semi-quantitative-PCR and fluorescence quantitative analysis, we found that Mlo gene showed the highest expression levels in leaves and the lowest in roots after inoculated with powdery mildew pathogen for different days.
文摘ESTs fragments which represents corresponding novel genes were obtained by sequencing and bioinformatics analysis of human fet al kidney cDNA library. Microarray was prepared by using these novel EST fragmen ts by automatic spotting. Expression patters of 79 ESTs of novel genes from huma n fetal kidney were analyzed in fetal brain and fetal heart tissues of 20\|week\ | and 26\|week\|age fetus by performing of cDNA chip hybridization. This provide s clues for studying exact functions of the novel genes. 8 genes were obtained w hich were expressed differentially in the fetal brain and heart of 20\|week\| an d 26\|week\|age respectively. Then differentially expressed genes were identifie d by Northern analysis. The more exact function of the novel genes is under stud y.
基金supported by grants from Wuhan Youth Chen-guang Project (No. 201050231077)National Natural Science Foundation of China (No. 30600571 and No. 81172786)
文摘In order to confirm the existence of indoleamine 2,3-dioxygenase(IDO) gene in swine,and to clone the novel gene followed by the molecule structure properties and expression pattern analysis,the porcine mRNA sequences homologous to human IDO were obtained from GenBank database by bioinformatics method.By using RT-PCR,the IDO gene was cloned from porcine endothelial cell line and the accuracy of the nucleic acid sequence was confirmed,and the expression pattern of the gene was detected.The three-dimensional structure model of porcine IDO was built referring to the tertiary structure of human IDO using biological sequence analysis software and database.The results showed that the porcine IDO was identified by sequencing.The nucleotide sequences were confirmed as a novel gene after submitted to Genbank.Porcine IDO was expressed in the lung,thymus,epididymis and anterior chamber with a basic level,however in peripheral blood mononuclear cells(PBMCs) the IDO gene was highly expressed.The three-dimensional structure model of porcine IDO was similar to that of human IDO.It was suggested that identification of the structure information of porcine IDO is essential to further investigate the immunologic function of the gene.Study of IDO on NK cells-mediated xenograft rejection will be a novel therapeutic target for the development of xenotransplantation.
基金supported by a grant from the National"863"High-Tech Program of China(Nos.102-10-01-05,Z19-01-01-03)the National"973"Key Basic Research Program of China(Fundamental Investigation on HumanCarcinogenesis)(No.G1998051008)and a grant from Chinese Medicine Board of New York,Inc.(No.96655)
文摘Objective: To identify the differential expression profile of human novel gene UBAP1, a putative nasopharyngeal neoplasms (NPC) relate gene, in multiple cancers. Methods: We first present an EST approach for electronic Northern in silico to analyse expression patterns of UBAP1 in tumor and normal tissues. Full length cDNA of UBAP1 gene was taken as a “probe” sequence, and a blastn search was performed against human EST Database. The Blastn report can be used to determine the fold differences between the pedigree ESTs in different libraries. Especially, the ESTs corresponding to UBAP1 present in fifteen tumor-derived libraries were compared against their normal counterpart to produce an electronic differential expression profile. Second, the distinct down-regulation of UBAP1 in meningioma, glioma, and colorectal tumor was confirmed by differentially RT-PCR analysis. Results: Database surveys indicated that UBAP1 gene was not only ubiquitously expressed in many normal tissues with various levels but also differentially expressed in different tumor tissues, especially down-regulated in multiple neoplastic tissues such as brain, breast, skin, colon, testis and uterus tumor tissues. Furthermore, differential RT-PCR analysis demonstrated that expression of UBAP1 was down-regulated or absent in 7 of 12 (58%) meningioma samples, 6 of 9 (66%) glioma and 7 of 11 (63%) colorectal tumor tissues respectively. Conclusion: we described a data mining procedure in silico that proved to be useful for the identification of differential expression patterns of UBAP1. These findings could be valuable for the investigation of the mechanism the differential expression of UBAP1 gene and its significance in the progression of multiple cancers.
文摘A repeated sequence DNA fragment, L5B-4, was cloned from the 5 kb BamHI DNA fragments of rat genomic DNA. The expressions of the L5B-4 DNA fragment are different in liver and hepatoma cells. The amounts of transcripts in hepatoma cells are lower in nucleus and higher in cytoplasm, especially in polysomal RNA, as compared with that in liver cells. The alteration shown in polysomal RNA of hepatoma cells seems to be specific. These results are discussed with respect to the possible function of this repeated DNA and its variation in hepatoma cells.
文摘Rhododendron simsii（Ericaeae：Rhododendron） has high ornamental value and ecological value.In this study,7 pairs of novel EST-SSR markers were developed from the genomic sequence of R.simsii,and they were used to investigate the genetic diversity of 32 natural R.simsii samples from Guifeng Mountain,Hubei Province.Results showed that a total of 31 polymorphic bands were amplified with allele number per locus of 4.43.Mean values of heterozygosity（Ho） and expected heterozygosity（He） were 0.679 58 and 0.723 14,respectively.This research will not only enrich the existing SSR database,but also lay a foundation for subsequent studies about molecular marker-assisted breeding,genetic diversity analysis,genetic structure analysis and phylogenetic analysis.
文摘Powdery mildew is a serious disease of wheat in China. As part of ITEC (International Triteace EST cooperation), EST (expressed sequence tags) technique was used to explore the gene expression in leaf induced by Erysiphe graminis DC. A conventional cDNA library was constructed, and a total of 1 500 clones picked randomly from the library were sequenced, three hundred and eighty_seven ESTs of them were unique, which got the Accession Number in GenBank. About 49.4% ESTs showed significant similarity to functions of known sequences in GenBank. There are 196 ESTs' with functions not able to be determined, and eighty_four ESTs were demonstrated to be novel sequences. High_density dot membranes from unique clones were produced, and several disease resistance related genes were screened by differential hybridization.
文摘Objective: To identify a cDNA clone from the subtracted library of human hepatocellular carcinoma (HCC). Methods: Suppression subtractive hybridization was used to isolated a panel of genes that are differentially expressed in hepatocellular carcinoma as compared with cirrhotic liver. T/A cloning method was used to construct a subtracted cDNA library. DNA sequencing analysis and Northern blot analysis were also utilized. Results: The cloned cDNA is 787 nucleotides in length and contains an open reading frame of 230 amino acids, which is a cDNA fragment of reported human fibrinogen, gamma polypeptide (FGG). Northern analysis revealed that this gene was overexpressed in two hepatocellular carcinoma cell lines, SMMC-7721 and HepG2. Conclusion: Sequence identity proved the cDNA clone fragment of as FGG gene. Differential expression of the cDNA fragment in HCC suggested that FGG is related to HCC, indicating a new clue for developing a novel diagnostic and prognostic marker.
文摘Seventy-five previously known plant microRNAs (miRNAs) were classified into 14 families according to their gene sequence identity. A total of 18,694 plant expressed sequence tags (EST) were found in the GenBank EST databases by comparing all previously known Arabidopsis miRNAs to GenBank’s plant EST databases with BLAST algorithms. After removing the EST sequences with high numbers (more than 2) of mismatched nucleotides, a total of 812 EST contigs were identified. After predicting and scoring the RNA secondary structure of the 812 EST sequences using mFold software, 338 new potential miRNAs were identified in 60 plant species. miRNAs are widespread. Some microRNAs may highly conserve in the plant kingdom, and they may have the same ancestor in very early evolution. There is no nucleotide substitution in most miRNAs among many plant species. Some of the new identified potential miRNAs may be induced and regulated by environmental biotic and abiotic stresses. Some may be preferentially expressed in specific tissues, and are regulated by developmental switching. These findings suggest that EST analysis is a good alternative strategy for identifying new miRNA candidates, their targets, and other genes. A large number of miRNAs exist in different plant species and play important roles in plant developmental switching and plant responses to environmental abiotic and biotic stresses as well as signal transduction. Environmental stresses and developmental switching may be the signals for synthesis and regulation of miRNAs in plants. A model for miRNA induction and expression, and gene regulation by miRNA is hypothesized.
基金Project supported by the National Natural Science Foundation of China(Nos.31101538,31000942,and 31000676)the Grand Science and Technology Special Project of Zhejiang Province(Nos.2010 C02006,2012C12903-4-1,and 2012C12903-6-3)+2 种基金the Zhejiang Provincial Natural Science Foundation of China(No.LY12C15004)the Public Welfare Project of Zhejiang Province(No.2011C22011)the Shaoxing Important Science and Technology Projects (No.2012A22008),China
文摘The development of expressed sequence tag-derived simple sequence repeats(EST-SSRs) provided a useful tool for investigating plant genetic diversity.In the present study,22 polymorphic EST-SSRs from grain soybean were identified and used to assess the genetic diversity in 48 vegetable soybean accessions.Among the 22 EST-SSR loci,tri-nucleotides were the most abundant repeats,accounting for 50.00% of the total motifs.GAA was the most common motif among tri-nucleotide repeats,with a frequency of 18.18%.Polymorphic analysis identified a total of 71 alleles,with an average of 3.23 per locus.The polymorphism information content(PIC) values ranged from 0.144 to 0.630,with a mean of 0.386.Observed heterozygosity(H o) values varied from 0.0196 to 1.0000,with an average of 0.6092,while the expected heterozygosity(H e) values ranged from 0.1502 to 0.6840,with a mean value of 0.4616.Principal coordinate analysis and phylogenetic tree analysis indicated that the accessions could be assigned to different groups based to a large extent on their geographic distribution,and most accessions from China were clustered into the same groups.These results suggest that Chinese vegetable soybean accessions have a narrow genetic base.The results of this study indicate that EST-SSRs from grain soybean have high transferability to vegetable soybean,and that these new markers would be helpful in taxonomy,molecular breeding,and comparative mapping studies of vegetable soybean in the future.
基金The Key Science and Technology Program of Shandong Province under contract No. 2012GHY11527Natural Science Foundation of Shandong Province under contract No. Q2007E02+1 种基金Specialized Research Fund for the Doctoral Program of Higher Education (New Teachers) under contract No. 20070423027the Public Science and Technology Research Funds Projects of Ocean, State Oceanic Administration of the People’s Republic of China under contract No. 201105021-8
文摘Zostera marina, a monocotyledonous angiosperm, is one of the most important seagrass species. To inves- tigate the salt-tolerance mechanism and discover salt-tolerant genes in Z. marina, a cDNA library was con- structed. Single-pass sequencing of the 5＇ ends of 4 081 clones yielded 4 002 high quality expressed sequence tags （ESTs）, which were assembled into 241 contigs and 1 673 singletons, representing 1 914 unigenes. The average length of the ESTs was 582 bp, with sizes ranging from 100-1 500 bp. Basic Local Alignment Search Tool （BLASTX） analysis revealed that 1 664 unigenes had significant homology to known genes in the Na- tional Center for Biotechnology Information （NCBI） non-redundant （nr） database （E-value≤5-10）. Among them, the two most abundant genes encoded metallothionein （157 ESTs） and chlorophyll a/b-binding pro- tein （38 ESTs）, accounting for 7.1% and 1.7% of the total ESTs, respectively. Using Kyoto Encyclopedia of Genes and Genomes （KEGG）, 1 462 unigenes were assigned to 1 161 pathways （E-value≤5-10）. A total of 938 unigenes were assigned Gene Ontology （GO） terms based on the GO hierarchy analysis, and InterProScan searches recognized 1 003 InterPro families. Three genes for metallothionein in Z. marina that belonged to Class II was identified. Results of this study will improve understanding of the molecular mechanisms of saline tolerance in Z. marina.
基金supported by the National Natural Science Foundation of China(81473310,31260075,31560085)
文摘Paris polyphylla Smith var.yunnanensis(Franch.) Hand.-Mazz.is a rhizomatous,herbaceous,perennial plant that has been used for more than a thousand years in traditional Chinese medicine.It is facing extinction due to overharvesting.Steroids are the major therapeutic components in Paris roots,the commercial value of which increases with age.To date,no genomic data on the species have been available.In this study,transcriptome analysis of an 8-year-old root and a 4-year-old root provided insight into the metabolic pathways that generate the steroids.Using Illumina sequencing technology,we generated a high-quality sequence and demonstrated de novo assembly and annotation of genes in the absence of prior genome information.Approximately 87,577 unique sequences,with an average length of 614 bases,were obtained from the root cells.Using bioinformatics methods,we annotated approximately 65.51% of the unique sequences by conducting a similarity search with known genes in the National Center for Biotechnology Information's non-redundant database.The unique transcripts were functionally classified using the Gene Ontology hierarchy and the Kyoto Encyclopedia of Genes and Genomes database.Of 3082 genes that were identified as significantly differentially expressed between roots of different ages,1518(49.25%) were upregulated and 1564(50.75%) were downregulated in the older root.Metabolic pathway analysis predicted that 25 unigenes were responsible for the biosynthesis of the saponins steroids.These data represent a valuable resource for future genomic studies on this endangered species and will be valuable for efforts to genetically engineer P.polyphylla and facilitate saponin-rich plant development.
基金supported by the Key Program for Science and Technology Development of Henan Province [122102310174]the Zoology Key Subject of Henan Province
文摘Objective： To screen and analyze key express sequence tags （ESTs） which were differentially displayed in every period of SD rats＇ primary hepatic carcinoma and reveal the molecular mechanism of carcinogenesis. Methods： Using diethylnitrosamine （DENA） as a cancerigenic agent, animal models with different phases of primary hepatic cancer were constructed in SD rats. Rats were respectively sacrificed at d 14, d 28, d 56, d 77, d 105 and d 112 after the rats received DENA by gavage, then the livers were harvested. One part of the livers was classified according to their pathological changes, while the other was reserved for molecular mechanism studies on hepatocarcinogenesis. The differentially expressed genes were isolated from both normal and morbid tissues by mRNA differential display technique （DDRT-PCR）. After the fragments were sequenced, bioinformatics were .used to analyze the results. Results： Twelve differentially expressed cDNA fragments were obtained. Nine fragments had the homology with known cDNA clones, especially EST-7 was similar to BN/SsNHsdMCW mitochondrion gene and the identity was 100% which suggested EST-7 may be the part of BN/SsNHsdMCW mitochondrion gene. In contrast, other three fragments （EST-1, EST-3 and EST-5） had extremely low identity to any genes registered in GENBANK databases. Conclusions： BN/SsNHsdMCW mitochondrion gene was expressed in different periods of hepatocarcinogenesis. Moreover, EST-I, EST-3 and EST-5 were suggested to contribute to the development of rat hepatocarcinogenesis, and thus may be candidates of new targets of oncogenes or cancer suppressor genes.
基金supported by the Zhejiang Provincial Science and Technology Program, China (2007C32013)the Natural Science Foundation of Zhejiang Province, China (Y3090660)
文摘Faba bean （Vicia faba L.）, one of the most important legumes in the world, evolved different types of cultivars due to its partial cross-pollination. The development of simple sequence repeat （SSR） markers from expressed sequence tags （EST） provided a useful tool for investigation of its genetic diversity. The purpose of the present study was to investigate the genetic diversity of faba bean from China and Europe using EST-SSR markers. 5 031 faba bean ESTs from the NCBI database were downloaded and assembled into 1 148 unigenes. A total of 107 microsatellites in 96 unigenes were identified, indicating that merely 8.36% of sequences contained SSRs. The most abundant SSR within faba bean was tri-nucleotide repeat motif, and among all the tri-nucleotide repeats, the motif AAG/CTT was the most abundant type. Based on these results, 11 EST-SSR markers were used to assess the genetic diversity of 29 faba bean cultivars from China and Europe with two to three alleles per locus. The polymorphism information content value ranged from 0.0644 to 0.4278 with an average of 0.2919. Principal coordinate analysis （PCA） and phylogenetic clustering based on these 11 EST-SSR markers distinguished these cultivars into different groups. The results indicated that faba bean in China had a narrow genetic basis, and the additional sources of genetic cultivars/accessions should be introduced to enhance the genetic variability. The results of this study proved that the EST-SSR marker is very effective in evaluation of faba bean germplasm.
基金Supported by the Scientific and Technical Supporting Programs of China (2008BAC49B01)the National Natural Science Foundation of China (No. 30830015)
文摘In 2008, a green tide broke out before the sailing competition of the 29th Olympic Games in Qingdao. The causative species was determined to be Enteromorpha prolifera （Ulva prolifera O. F. Miiller）, a familiar green macroalga along the coastline of China. Rapid accumulation of a large biomass of floating U. prolifera prompted research on different aspects of this species. In this study, we constructed a nonnormalized cDNA library from the thalli of U. prolifera and acquired 10072 high-quality expressed sequence tags （ESTs）. These ESTs were assembled into 3 519 nonredundant gene groups, including 1 446 clusters and 2 073 singletons. After annotation with the nr database, a large number of genes were found to be related with chloroplast and ribosomal protein, GO functional classification showed 1 418 ESTs participated in photosynthesis and 1 359 ESTs were responsible for the generation of precursor metabolites and energy. In addition, rather comprehensive carbon fixation pathways were found in U. prolifera using KEGG. Some stress-related and signal transduction-related genes were also found in this study. All the evidences displayed that U. prolifera had substance and energy foundation for the intense photosynthesis and the rapid proliferation. Phylogenetic analysis of cytochrome c oxidase subunit I revealed that this green-tide causative species is most closely affiliated to Pseudendoclonium akinetum （Ulvophyceae）.
基金Project (No. 30070070) supported by the National NaturalScience Foundation of China
文摘Aluminum (Al) toxicity is the major factor limiting crop productivity in acid soils. In this study, a recombinant inbreed line (RIL) population derived from a cross between an Al sensitive lowland indica rice variety IR1552 and an Al tolerant upland japonica rice variety Azucena, was used for mapping quantitative trait loci (QTLs) for Al tolerance. Three QTLs for relative root length (RRL) were detected on chromosome 1, 9, 12, respectively, and 1 QTL for root length under Al stress is identical on chromosome 1 after one week and two weeks stress. Comparison of QTLs on chromosome 1 from different studies indicated an identical interval between C86 and RZ801 with gene(s) for Al tolerance. This interval provides an important start point for isolating genes responsible for Al tolerance and understanding the genetic nature of Al tolerance in rice. Four Al induced ESTs located in this interval were screened by reverse Northern analysis and confirmed by Northern analysis. They would be candidate genes for the QTL.
文摘MicroRNAs(miRNA)are recently discovered endogenous,small noncoding RNAs(of 22 nucleotides)that play pivotal roles in gene regulation.They are involved in post-transcriptional control of gene expression.miRNAs are emerging as important regulators of cell proliferation,development,cancer formation,stress responses,cell death and physiological conditions.Increasing evidence has demonstrated the human miRNAs bind to their target mRNA sequences with perfect or near-perfect sequence complementarily.This provides a powerful strategy for discovering potential type 2 diabetes mellitus(T2DM)targets and gives the probability to exploit them for diagnostic and therapeutic causes.About 6%of the world population is affected by T2DM,and it is recognized as a global epidemic by the World Health Organization.At present there is no valid biomarker to control or manage T2DM.Therefore,the present study applied a mature sequence of miRNAs from publicly accessible databases to identify the miRNA from T2DM expressed sequence tags,and the results are detailed and discussed below.
基金supported by the Core Research Budget of the Non-Profit Governmental Research Institution(ICS,CAAS)(0032007216 and 201011)the National Natural Science Foundation of China(30800752)the Key Laboratory of Vegetable Genetics and Physiology,Ministry of Agriculture,China
文摘SSR markers are useful in pepper linkage mapping and gene location.446 SSR markers have been reported,but they are insufficient.It is costly to develop SSR markers from DNA library,whereas it seems much easy to find in EST sequences in the GenBank of pepper through internet.In this study,attempts have been made to develop SSR markers in the EST sequences by using bioinformatics.EST sequences were trimmed by‘est-trimmer.pl’software,while 116915 EST sequences were obtained without poly‘A’or poly‘T’,ranged between 100 and 700 bp.Using‘e-PCR’and‘del.pl’softwares,SSR sequences were identified.2508 microsatellite loci(larger than 20 repeats)were established and 755 SSR primers were designed using SSR finder software and Primer 3 software.There were 498(0.43%)mono-,1026(0.89%)di-,518(0.45%)tri-,245(0.21%)tetra-,114(0.10%)penta-,and 107(0.09%)hexa-nucleotide SSRs.The estimated frequency of SSRs was approximately 1/25.12 kb.According to the distribution of SSRs in pepper,the mean length of pepper SSRs was 22.68 bp and the adenine rich repeats such as A/T,AG,AT,AAG,AAAT,and AAAC were predominant in each type of SSRs(mono-,di-,tri-,tetra-,penta-,and hexa-),whereas the C/G,CG,CCG repeats were less abundant.210 primers were tested in 8 pepper cultivars and the PCR result revealed the existence of polymorphism among 127(60.48%)SSR primers within 8 pepper cultivars.It is confirmed that pepper EST database could be efficiently exploited for availability of SSR markers.
