The rapidly increasing number of sequences entering into the genome databank has created the need for fully automated methods to analyze them.Knowing the cellular location of a protein is a key step towards understand...The rapidly increasing number of sequences entering into the genome databank has created the need for fully automated methods to analyze them.Knowing the cellular location of a protein is a key step towards understanding its function.The development in statistical prediction of protein attributes generally consists of two cores: one is to construct a training dataset and the other is to formulate a predictive algorithm.The latter can be further separated into two subcores: one is how to give a mathematical expression to effectively represent a protein and the other is how to find a powerful algorithm to accurately perform the prediction.To predict the subcellular location of eukaryotic protein,a systematic prediction approach comprised of a novel feature extraction method,an idea of combining this feature extraction method with support vector machine(SVM) algorithm,and ’one-versus-rest’ & ’all-versus-all’ strategies have been proposed in this paper.Consequently,the total predictive accuracies reach 95.5% for four locations.Compared with existing methods,this new approach provides better predictive performance.For example,it is 13.5%,5.1% higher than Yuan’s and Hua’s methods respectively.These results demonstrate the applicability of this new method and concept and possible improvement of prediction for the protein subcellular location.It is anticipated that the current approach may also have a series of impacts on the prediction of other protein features.展开更多
Objective： To construct eukaryotic expression vector containing B7-1/GFP geneand study its expression in osteosarcoma cell line LM8. Methods： By using gene cloning technique, eukaxyotic expression vector pEGFP-C1 wa...Objective： To construct eukaryotic expression vector containing B7-1/GFP geneand study its expression in osteosarcoma cell line LM8. Methods： By using gene cloning technique, eukaxyotic expression vector pEGFP-C1 was used to construct the murine B7-1 recombinant plasmid （pEGFP-C1/B7）. Recombinant plasmid was transfected into LM8 cells with liposome and was confirmed by restriction endonuclease digestion and DNA sequencing. The expression of the fusion protein was detected using fluorescence microscope and Western blot analysis. Results： The recombinant eukaryotic expression plasmid pEGFP-C1/B7 was successfully constructed, which was confirmed by DNA sequencing, RT-PGR and restriction enzymes analysis. The green fluorescent protein could be detected in the transfected LM8 with fluorescence microscope. The expected B7-1 and green fluorescent protein （GFP） fusion protein was detected by RT-PCR and Western blot. Conclusion： The eukaryotic expression vector containing B7-1/GFP gene was constructed successfully, and it could be expressed in LM8 after transfection.展开更多
A new system is developed to recognize promoter sequences from non promoter sequences based on position weight matrix and backpropagation neural network in this paper. The system performs significantly better on the t...A new system is developed to recognize promoter sequences from non promoter sequences based on position weight matrix and backpropagation neural network in this paper. The system performs significantly better on the training set and the test set, the mean recognition rate is as high as 99% on the training set and 97% on the testing set. Experimental results demonstrate the effectiveness of the system to recognize the promoter sequences that have been trained and the promoter sequences that have not been seen previously.展开更多
Objective： To clone and construct eukaryotic expressing vectors of sense and antisense human Pin1 （hPinl） genes. Methods： Total RNA was extracted from MG-63 cells, then the hPinl cDNA was amplified by RT-PCR. The ...Objective： To clone and construct eukaryotic expressing vectors of sense and antisense human Pin1 （hPinl） genes. Methods： Total RNA was extracted from MG-63 cells, then the hPinl cDNA was amplified by RT-PCR. The same time the sense and antisense hPinl genes were formed by binding BamH Ⅰ and Hind Ⅲ in cis and trans-directions. At the end they were cloned into the eukaryotic expressing vector pIRES2-EGFP in cis and trans directions using DNA recombinant technology. The recombinant vectors were further identified by digestion of BamHⅠ and Hind Ⅲ. Results： The results of sequencing showed that the orientation of the ligations and the reading frame were correct. After digested by BamH Ⅰ and Hind Ⅲ, two fragments exhibiting 5.3 kb and 0.99 kb were formed in sense and antisense eukaryotic expressing vectors. Electrophoretic results were completely coincident with theoretical calculation. Conclusion： Human Pin1 sense and antisense genes were successfully cloned and eukaryotic expressing vectors were successfully constructed.展开更多
Nitzschia closterium f. minutissima, a marine eukaryotic unicellular diatom, originally classified as Bacillariophyta/Bacillariophyceae/Bacillariales/Bacillariaceae/Nitzschia, is one of the most important feed sources...Nitzschia closterium f. minutissima, a marine eukaryotic unicellular diatom, originally classified as Bacillariophyta/Bacillariophyceae/Bacillariales/Bacillariaceae/Nitzschia, is one of the most important feed sources in mariculture. In this study, its morphological features were examined under DIC Microscopy (differential interference contrast microscope); its pigments and fatty acids composition were analyzed by using High Performance Liquid Chromatography (HPLC) and gas chromatography (GC); the complete Actin cDNA, part 18S rDNA, complete ITS1 and ITS2 sequences, part 28S rDNA se- quences, and a putatively encoding ?5 fatty acid desaturase gene were cloned respectively and further functioned in transgenic yeast. The sequence alignments were separately conducted using the related sequences from Nitzschia closterium f. minutissima, Cylindrotheca closterium (Bacillariophyta/Baci- llariales/Bacillariaceae/Cylindrotheca) and Phaeodactylum tricornutum (Naviculales) with ClustalX 1.83. No distinct difference was discovered between N. closterium f. minutissima and P. tricornutum in both biochemical and molecular level. Their identity was more than 99.6% among 18S rDNA, 5.8S rDNA and actin-gene sequences, and is up to 98.6% even among ITS1 and ITS2 sequences. Their ?5 desaturase similarity was 99.4%. However, the lower similarity was disclosured between N. closterium f. minutis- sima and Cylindrotheca closterium, which shared less than 40% identity in the ITS1 and ITS2 se- quences. So, N. closterium f. minutissima should not be placed in Bacillariales, Bacillariaceae, Nitzschia, but in Naviculales, Phaeodactylaceae, Phaeodactylum, and it was actually a strain of P. tri- cornutum.展开更多
文摘The rapidly increasing number of sequences entering into the genome databank has created the need for fully automated methods to analyze them.Knowing the cellular location of a protein is a key step towards understanding its function.The development in statistical prediction of protein attributes generally consists of two cores: one is to construct a training dataset and the other is to formulate a predictive algorithm.The latter can be further separated into two subcores: one is how to give a mathematical expression to effectively represent a protein and the other is how to find a powerful algorithm to accurately perform the prediction.To predict the subcellular location of eukaryotic protein,a systematic prediction approach comprised of a novel feature extraction method,an idea of combining this feature extraction method with support vector machine(SVM) algorithm,and ’one-versus-rest’ & ’all-versus-all’ strategies have been proposed in this paper.Consequently,the total predictive accuracies reach 95.5% for four locations.Compared with existing methods,this new approach provides better predictive performance.For example,it is 13.5%,5.1% higher than Yuan’s and Hua’s methods respectively.These results demonstrate the applicability of this new method and concept and possible improvement of prediction for the protein subcellular location.It is anticipated that the current approach may also have a series of impacts on the prediction of other protein features.
基金This study was supported by a grant from the youth dawn program of Wuhan (Grant No. 20025001028).
文摘Objective： To construct eukaryotic expression vector containing B7-1/GFP geneand study its expression in osteosarcoma cell line LM8. Methods： By using gene cloning technique, eukaxyotic expression vector pEGFP-C1 was used to construct the murine B7-1 recombinant plasmid （pEGFP-C1/B7）. Recombinant plasmid was transfected into LM8 cells with liposome and was confirmed by restriction endonuclease digestion and DNA sequencing. The expression of the fusion protein was detected using fluorescence microscope and Western blot analysis. Results： The recombinant eukaryotic expression plasmid pEGFP-C1/B7 was successfully constructed, which was confirmed by DNA sequencing, RT-PGR and restriction enzymes analysis. The green fluorescent protein could be detected in the transfected LM8 with fluorescence microscope. The expected B7-1 and green fluorescent protein （GFP） fusion protein was detected by RT-PCR and Western blot. Conclusion： The eukaryotic expression vector containing B7-1/GFP gene was constructed successfully, and it could be expressed in LM8 after transfection.
文摘A new system is developed to recognize promoter sequences from non promoter sequences based on position weight matrix and backpropagation neural network in this paper. The system performs significantly better on the training set and the test set, the mean recognition rate is as high as 99% on the training set and 97% on the testing set. Experimental results demonstrate the effectiveness of the system to recognize the promoter sequences that have been trained and the promoter sequences that have not been seen previously.
文摘Objective： To clone and construct eukaryotic expressing vectors of sense and antisense human Pin1 （hPinl） genes. Methods： Total RNA was extracted from MG-63 cells, then the hPinl cDNA was amplified by RT-PCR. The same time the sense and antisense hPinl genes were formed by binding BamH Ⅰ and Hind Ⅲ in cis and trans-directions. At the end they were cloned into the eukaryotic expressing vector pIRES2-EGFP in cis and trans directions using DNA recombinant technology. The recombinant vectors were further identified by digestion of BamHⅠ and Hind Ⅲ. Results： The results of sequencing showed that the orientation of the ligations and the reading frame were correct. After digested by BamH Ⅰ and Hind Ⅲ, two fragments exhibiting 5.3 kb and 0.99 kb were formed in sense and antisense eukaryotic expressing vectors. Electrophoretic results were completely coincident with theoretical calculation. Conclusion： Human Pin1 sense and antisense genes were successfully cloned and eukaryotic expressing vectors were successfully constructed.
基金the National Basic Research Program of China (Grant No. 2005CCA02400)
文摘Nitzschia closterium f. minutissima, a marine eukaryotic unicellular diatom, originally classified as Bacillariophyta/Bacillariophyceae/Bacillariales/Bacillariaceae/Nitzschia, is one of the most important feed sources in mariculture. In this study, its morphological features were examined under DIC Microscopy (differential interference contrast microscope); its pigments and fatty acids composition were analyzed by using High Performance Liquid Chromatography (HPLC) and gas chromatography (GC); the complete Actin cDNA, part 18S rDNA, complete ITS1 and ITS2 sequences, part 28S rDNA se- quences, and a putatively encoding ?5 fatty acid desaturase gene were cloned respectively and further functioned in transgenic yeast. The sequence alignments were separately conducted using the related sequences from Nitzschia closterium f. minutissima, Cylindrotheca closterium (Bacillariophyta/Baci- llariales/Bacillariaceae/Cylindrotheca) and Phaeodactylum tricornutum (Naviculales) with ClustalX 1.83. No distinct difference was discovered between N. closterium f. minutissima and P. tricornutum in both biochemical and molecular level. Their identity was more than 99.6% among 18S rDNA, 5.8S rDNA and actin-gene sequences, and is up to 98.6% even among ITS1 and ITS2 sequences. Their ?5 desaturase similarity was 99.4%. However, the lower similarity was disclosured between N. closterium f. minutis- sima and Cylindrotheca closterium, which shared less than 40% identity in the ITS1 and ITS2 se- quences. So, N. closterium f. minutissima should not be placed in Bacillariales, Bacillariaceae, Nitzschia, but in Naviculales, Phaeodactylaceae, Phaeodactylum, and it was actually a strain of P. tri- cornutum.
