Insect pheromone-binding proteins (PBPs) play important roles in transporting hydrophobic pheromone components across the sensillum lymph to the surface of olfactory receptors (ORs). However, the PBPs of the orien...Insect pheromone-binding proteins (PBPs) play important roles in transporting hydrophobic pheromone components across the sensillum lymph to the surface of olfactory receptors (ORs). However, the PBPs of the oriental fruit moth, Grapholita molesta, an important destructive pest of stone fruits worldwide, are not well characterized. In this study, two new putative PBP genes, GmolPBP2 and GmolPBP3, were identiifed from G. molesta antennae. The deduced amino-acid sequences of these two putative PBP genes are characteristic of the odorant binding protein family, containing six conserved cysteine residues. The genomic DNA sequence of each gene contained two introns. However, the lengths and positions of the introns differed. RT-PCR analyses revealed that the two GmolPBP genes are only expressed in the antennae of female and male moths. Quantitative real-time PCR indicated that the transcription levels of GmolPBP2 are far greater than those of GmolPBP3 in both female and male antennae. GmolPBP3 showed higher transcription levels in female antennae than in male antennae, while GmolPBP2 showed similar transcription levels in both female and male antennae. The transcript levels of both genes were signiifcantly different in premating and post-coitum individuals, implying that mating affects the process of sex pheromone reception. To better understand the functions, two GmolPBPs were expressed in Escherichia coli, and the ligand binding assays were conducted. Results showed that GmolPBP2 has strong binding afifnities to two sex pheromone components, E8-12:Ac and Z8-12:Ac, as well as weaker binding afifnities to Z8-12:OH and 12:OH. GmolPBP2 also bound some ordinary odor molecules. However, the afifnity of GmolPBP3 to both sex pheromones and ordinary odor molecules was very weak. These results show that GmolPBP2 plays the main role in pheromone discrimination and recognition in the oriental fruit moth.展开更多
[ Objective] This study aimed to investigate the optimal induction conditions for expression of recombinant fusion protein. [ Method] The optimal ini- tial OD600, induction dose, induction duration, induction temperat...[ Objective] This study aimed to investigate the optimal induction conditions for expression of recombinant fusion protein. [ Method] The optimal ini- tial OD600, induction dose, induction duration, induction temperature, ampicillin concentration, liquid volume and other culture conditions were explored by using parallel fermentation experiments with different technical indicators, to improve the expression level of the recombinant fusion protein of IMPACT-CN pTYB11 vector. [Result] SDS-PAGE and western-blotting analysis show that the recombinant fusion protein has a high immunogenicity. [Conclusion] This study laid the foundation for further protein purification in diagnosis, production of diagnostic kits with protein and clinical application.展开更多
CAP,an adenylyl cyclase-associated protein,is predicted to be involved in cytoskeletal organization and signal transduction.Recently,we found that CAP may play an important role in fuzz-like fiber cell initiation in c...CAP,an adenylyl cyclase-associated protein,is predicted to be involved in cytoskeletal organization and signal transduction.Recently,we found that CAP may play an important role in fuzz-like fiber cell initiation in cotton.For the further research,we isolated two CAP homologues from wild展开更多
The zinc finger proteins belong to the largest family of transcription factors.But there is little research of Cys2/His2 type zinc finger proteins in cotton,and there is no submission of correlating
Acyl-ACP thioesterases (FATs) terminates the fatty acid synthesis and allow the transport of fatty acids out of the plastids, which are the important determinants of cellular metabolism. FATB is a member of FAT enzy...Acyl-ACP thioesterases (FATs) terminates the fatty acid synthesis and allow the transport of fatty acids out of the plastids, which are the important determinants of cellular metabolism. FATB is a member of FAT enzymes that has been described previously in most of the plants. In silico cloning is a new method that utilizes the bioinformatics on the complete genome and available EST database. In this study, a full-length cDNA clone of PtFATB gene was isolated from Populus tomentosa using this approach. It is 1,450 bp in length and the open reading frame encodes a peptide of 421 amino acids. The predicted amino acid sequence shows significant homology with those from other plant species, which contain typical domains owned by FATB proteins. The transcripts of PtFATB were abundant in leaves, and less in roots detected by using semiquantitative RT-PCR. When the shoots were subjected to the stress treatments (cold, dry, NaC1) and ABA (Abscisic acid), the expression of PtFATB was only slightly reduced under the treatment of low temperature. This suggests that the expression of PtFATB is in a constitutive fashion. This study provides the basis not only for the identification and characterization of this gene but also for the improvement of cold tolerance by controlling the expression of the PtFATB gene in trees in near future.展开更多
In higher plants, WRKY gene family plays a significant role in transcriptional regulation. They are widely involved in biotic stress, abiotic stress, growth, development and metabolism regulation. In this study WRKY6 ...In higher plants, WRKY gene family plays a significant role in transcriptional regulation. They are widely involved in biotic stress, abiotic stress, growth, development and metabolism regulation. In this study WRKY6 from tobacco is cloned by homology cloning. 1647 nucleotide sequences were obtained. The deduced protein sequences show that this protein belongs to the second group of WRKY family, only have one WRKY structure, and the zinc-finger structure is C-X4-C-X23-H-X1-H. Phylogeny results show NtWRKY6 is much closer to NtWRKY1 generated 97% amino acids similarity. RT-PCR analysis has revealed that expression levels of NtWRKY6 has increased rapidly at 3 h under NaCI and PEG treatment. The results suggest that NtWRKY6 is an early responder and may be involved in NaCl and PEG abiotic stress in tobacco.展开更多
AIM: Using bacterial, yeast, or mammalian cell expressing a human drug metabolism enzyme would seem good way to study drug metabolism-related problems. Human cytochrome P-450 2C9(CYP2C9) is a polymorphic enzyme respon...AIM: Using bacterial, yeast, or mammalian cell expressing a human drug metabolism enzyme would seem good way to study drug metabolism-related problems. Human cytochrome P-450 2C9(CYP2C9) is a polymorphic enzyme responsible for the metabolism of a large number of clinically important drugs. It ranks among the most important drug metabolizing enzymes in humans. In order to provide a sufficient amount of the enzyme for drug metabolic research, the CYP2C9 cDNA was cloned and expressed stably in CHL cells. METHODS: After extraction of total RNA from human liver tissue, the human CYP2C9 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR), and cloned into cloning vector pGEM-T. The cDNA fragment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9. A transgenic cell line was established by transfecting the recombinant vector of pREP9-CYP2C9 into CHL cells. The enzyme activity of CYP2C9 catalyzing oxidation of tolbutamide to hydroxy tolbutamide in S9 fraction of the cell was determined by high performance liquid chromatography(HPLC). RESULTS: The amino acid sequence predicted from the cDNA segment was identical to that of CYP2C9*1, the wild type CYP2C9. However, there were two base differences, i.e. 21T】C, 1146C】T, but the encoding amino acid sequence was the same, L7, P382. The S9 fraction of the established cell line metabolizes tolbutamide to hydroxy tolbutamide; tolbutamide hydroxylase activity was found to be 0.465 +/- 0.109 micromol.min(-1).g(-1) S9 protein or 8.62 +/- 2.02mol.min(-1).mol(-1) CYP, but was undetectable in parental CHL cell. CONCLUSION: The cDNA of human CYP2C9 was successfully cloned and a cell line of CHL- CYP2C9, efficiently expressing the protein of CYP2C9, was established.展开更多
Bicoid is one of the important Drosophila maternal genes involved in the control of embryo polarity and larvae segmentation. To clone and characterize the rice bicoid-related genes, one cDNA clone, Rb24 (EMBL accessio...Bicoid is one of the important Drosophila maternal genes involved in the control of embryo polarity and larvae segmentation. To clone and characterize the rice bicoid-related genes, one cDNA clone, Rb24 (EMBL accession number: AJ2771380), was isolated by screening of rice unmature seed cDNA library. Sequence analysis indicates that Rb24 contains a putative amino acid sequence, which is homologous to unique 8 amino acids sequence within Drosophila bicoid homeodomain (50% identity, 75% similarity) and involves a lys-9 in putative helix 3. Northern blot analysis of rice RNA has shown that this sequence is expressed in a tissue-specific manner. The transcript was detected strongly in young panicles, but less in young leaves and roots. This results are further confirmed with paraffin section in situ hybridization. The signal is intensive in rice globular embryo and located at the apical tip of the embryo, then, along with the development of embryo, the signal is getting reduced and transfers into both sides of embryo. The existence of bicoid-related sequence in rice embryo and the similarity of polar distribution of bicoid and Rb24 mRNA in early embryo development may implicates a conserved maternal regulation mechanism of body axis presents in Drosophila and in rice.展开更多
Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules...Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules. Despite extensive research on nsLTP in different plant species, mostly angiosperm, and the great diversity of physiological processes in which they seem to be involved, their exact functions still remain unclear. Also, very limited experimental data are available on nsLTP in gymnosperm. In this study, we report for the first time on the molecular cloning of nsLTP, from Pinus sylvestris L.(PsLTP1, GenBank accession JN980402.1) and the expression pattern of PsLTP1 during ontogenesis and in response to environmental stress conditions. Total RNA from roots of 7-day old pine seedlings was used to isolate the cDNA clone, corresponding to Scots pine lipid transfer protein. The open reading frame of PsLTP1 consists of 372 bp encoding a protein of 123 amino acids. Amino acid sequence alignment revealed that mature PsLTP1 shares high level of similarity with nsLTP from other conifers and with well-studied nsLTPs from angiosperms. The PsLTP1 contains a 27-amino-acid N-terminal signal sequence and presents all the features of a plant nsLTP. Amino acid comparison analysis and 3D structure prediction showed that PsLTP1 is a type 1 nsLTP. The results of the expression analysis of Scots pine PsLTP1 gene revealed that its transcripts accumulate in actively growing tissues. Furthermore, transcription of PsLTP1 was upregulated in response to cold and salt treatments, and downregulated during acidic, osmotic and water stresses.展开更多
AIM: To develop and optimize cDNA representational difference analysis (cDNA RDA) method and to identify and clone garlic up-regulated genes in human gastric cancer (HGC) cells. METHODS: We performed cDNA RDA method b...AIM: To develop and optimize cDNA representational difference analysis (cDNA RDA) method and to identify and clone garlic up-regulated genes in human gastric cancer (HGC) cells. METHODS: We performed cDNA RDA method by using abundant double-stranded cDNA messages provided by two self-constructed cDNA libraries (Allitridi-treated and paternal HGC cell line BGC823 cells cDNA libraries respectively). Bam H I and Xho I restriction sites harbored in the library vector were used to select representations. Northern and Slot blots analyses were employed to identify the obtained difference products. RESULTS: Fragments released from the cDNA library vector after restriction endonuclease digestion acted as good marker indicating the appropriate digestion degree for library DNA. Two novel expressed sequence tags (ESTs) and a recombinant gene were obtained. Slot blots result showed a 8-fold increase of glia-derived nexin/protease nexin 1 (GDN/PN1) gene expression level and 4-fold increase of hepatitis B virus x-interacting protein (XIP) mRNA level in BGC823 cells after Allitridi treatment for 72h. CONCLUSION: Elevated levels of GDN/PN1 and XIP mRNAs induced by Allitridi provide valuable molecular evidence for elucidating the garlic's efficacies against neurodegenerative and inflammatory diseases. Isolation of a recombinant gene and two novel ESTs further show cDNA RDA based on cDNA libraries to be a powerful method with high specificity and reproducibility in cloning differentially expressed genes.展开更多
Chemosensory proteins (CSPs) are ubiquitous soluble small proteins isolated from sensory organs thought to be involved in chemical communication. Here we report the first cDNA of CSPs, called Ac-ASP3, cloned and chara...Chemosensory proteins (CSPs) are ubiquitous soluble small proteins isolated from sensory organs thought to be involved in chemical communication. Here we report the first cDNA of CSPs, called Ac-ASP3, cloned and characterized from antennas of adult worker bees in Chinese honeybee, Apis cerana cerana. The Ac-ASP3 cDNA comprises 2 exons, with an ORF of 393-bp encoding 130 aa. Protein signature analyses show that the protein consists of four conserved cysteines and a signal peptide with 19 aa in the N-terminal sequence. The deduced protein sequence shares high homology with Am-ASP3 of Apis mellifera and low similarity with other species of insects. Immunocytochemical localization shows that Ac-ASP3 is only specifically expressed on the antenna contact chemosensilla such as sensilla trichodea B and sensilla basiconica, whereas Ac-ASP3 is scarcely expressed on olfactory chemosensilla such as sensilla placodea. Real-time PCR of Ac-ASP3 transcripts shows that Ac-ASP3 is highly expressed on wings and legs, but expression is lower on antenna. Temporal expression patterns suggest that Ac-ASP3 is expressed during the period of pupa and adults from 1-d to 6-d stages when bees act as house bees, cleaning the comb and taking care of the queen and larvae in comb. The above evidence suggests that Ac-ASP3 is unique in species and is generally not involved in olfaction during searching for honey and pollen. Rather, the protein seems to function in recognition of chemosensory substances on bees' cuticle and mechanical movement of antenna.展开更多
The large protein of the hepatitis B virus (HBV) envelope was subdivided intothe preSl region with 108-115 amino acids, the preS2 region with 55 amino acidsand the S region with 226 amino acids. It was believed that t...The large protein of the hepatitis B virus (HBV) envelope was subdivided intothe preSl region with 108-115 amino acids, the preS2 region with 55 amino acidsand the S region with 226 amino acids. It was believed that the preSl region cotainedthe hepatocyte attachment sites of the virus as well as multiple highly immunogenicT-cell and B-cell epitopes. The antibody response to the preSl peptide was also oneof the most important serological markers of HBV infection.展开更多
基金supported by the National Natural Science Foundation of China (31272043)the the Special Fund for Agro-Scientific Research in the Public Interest, China (201103024)
文摘Insect pheromone-binding proteins (PBPs) play important roles in transporting hydrophobic pheromone components across the sensillum lymph to the surface of olfactory receptors (ORs). However, the PBPs of the oriental fruit moth, Grapholita molesta, an important destructive pest of stone fruits worldwide, are not well characterized. In this study, two new putative PBP genes, GmolPBP2 and GmolPBP3, were identiifed from G. molesta antennae. The deduced amino-acid sequences of these two putative PBP genes are characteristic of the odorant binding protein family, containing six conserved cysteine residues. The genomic DNA sequence of each gene contained two introns. However, the lengths and positions of the introns differed. RT-PCR analyses revealed that the two GmolPBP genes are only expressed in the antennae of female and male moths. Quantitative real-time PCR indicated that the transcription levels of GmolPBP2 are far greater than those of GmolPBP3 in both female and male antennae. GmolPBP3 showed higher transcription levels in female antennae than in male antennae, while GmolPBP2 showed similar transcription levels in both female and male antennae. The transcript levels of both genes were signiifcantly different in premating and post-coitum individuals, implying that mating affects the process of sex pheromone reception. To better understand the functions, two GmolPBPs were expressed in Escherichia coli, and the ligand binding assays were conducted. Results showed that GmolPBP2 has strong binding afifnities to two sex pheromone components, E8-12:Ac and Z8-12:Ac, as well as weaker binding afifnities to Z8-12:OH and 12:OH. GmolPBP2 also bound some ordinary odor molecules. However, the afifnity of GmolPBP3 to both sex pheromones and ordinary odor molecules was very weak. These results show that GmolPBP2 plays the main role in pheromone discrimination and recognition in the oriental fruit moth.
基金Supported by Binzhou Polytechnic Scientific Research Project ( 07XYA04)
文摘[ Objective] This study aimed to investigate the optimal induction conditions for expression of recombinant fusion protein. [ Method] The optimal ini- tial OD600, induction dose, induction duration, induction temperature, ampicillin concentration, liquid volume and other culture conditions were explored by using parallel fermentation experiments with different technical indicators, to improve the expression level of the recombinant fusion protein of IMPACT-CN pTYB11 vector. [Result] SDS-PAGE and western-blotting analysis show that the recombinant fusion protein has a high immunogenicity. [Conclusion] This study laid the foundation for further protein purification in diagnosis, production of diagnostic kits with protein and clinical application.
文摘CAP,an adenylyl cyclase-associated protein,is predicted to be involved in cytoskeletal organization and signal transduction.Recently,we found that CAP may play an important role in fuzz-like fiber cell initiation in cotton.For the further research,we isolated two CAP homologues from wild
文摘The zinc finger proteins belong to the largest family of transcription factors.But there is little research of Cys2/His2 type zinc finger proteins in cotton,and there is no submission of correlating
基金This work was supported by project "Regulation of Composition and Saturation of Fatty Acid in Trees by Genetic Engineering", Introduction of Foreign Advanced Agricultural Science and Technology into China (No. 2005-4-52).
文摘Acyl-ACP thioesterases (FATs) terminates the fatty acid synthesis and allow the transport of fatty acids out of the plastids, which are the important determinants of cellular metabolism. FATB is a member of FAT enzymes that has been described previously in most of the plants. In silico cloning is a new method that utilizes the bioinformatics on the complete genome and available EST database. In this study, a full-length cDNA clone of PtFATB gene was isolated from Populus tomentosa using this approach. It is 1,450 bp in length and the open reading frame encodes a peptide of 421 amino acids. The predicted amino acid sequence shows significant homology with those from other plant species, which contain typical domains owned by FATB proteins. The transcripts of PtFATB were abundant in leaves, and less in roots detected by using semiquantitative RT-PCR. When the shoots were subjected to the stress treatments (cold, dry, NaC1) and ABA (Abscisic acid), the expression of PtFATB was only slightly reduced under the treatment of low temperature. This suggests that the expression of PtFATB is in a constitutive fashion. This study provides the basis not only for the identification and characterization of this gene but also for the improvement of cold tolerance by controlling the expression of the PtFATB gene in trees in near future.
文摘In higher plants, WRKY gene family plays a significant role in transcriptional regulation. They are widely involved in biotic stress, abiotic stress, growth, development and metabolism regulation. In this study WRKY6 from tobacco is cloned by homology cloning. 1647 nucleotide sequences were obtained. The deduced protein sequences show that this protein belongs to the second group of WRKY family, only have one WRKY structure, and the zinc-finger structure is C-X4-C-X23-H-X1-H. Phylogeny results show NtWRKY6 is much closer to NtWRKY1 generated 97% amino acids similarity. RT-PCR analysis has revealed that expression levels of NtWRKY6 has increased rapidly at 3 h under NaCI and PEG treatment. The results suggest that NtWRKY6 is an early responder and may be involved in NaCl and PEG abiotic stress in tobacco.
基金National Natural Science Foundation of China,No.39770868Natural Science Foundation of Zhejiang Province,No.397490
文摘AIM: Using bacterial, yeast, or mammalian cell expressing a human drug metabolism enzyme would seem good way to study drug metabolism-related problems. Human cytochrome P-450 2C9(CYP2C9) is a polymorphic enzyme responsible for the metabolism of a large number of clinically important drugs. It ranks among the most important drug metabolizing enzymes in humans. In order to provide a sufficient amount of the enzyme for drug metabolic research, the CYP2C9 cDNA was cloned and expressed stably in CHL cells. METHODS: After extraction of total RNA from human liver tissue, the human CYP2C9 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR), and cloned into cloning vector pGEM-T. The cDNA fragment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9. A transgenic cell line was established by transfecting the recombinant vector of pREP9-CYP2C9 into CHL cells. The enzyme activity of CYP2C9 catalyzing oxidation of tolbutamide to hydroxy tolbutamide in S9 fraction of the cell was determined by high performance liquid chromatography(HPLC). RESULTS: The amino acid sequence predicted from the cDNA segment was identical to that of CYP2C9*1, the wild type CYP2C9. However, there were two base differences, i.e. 21T】C, 1146C】T, but the encoding amino acid sequence was the same, L7, P382. The S9 fraction of the established cell line metabolizes tolbutamide to hydroxy tolbutamide; tolbutamide hydroxylase activity was found to be 0.465 +/- 0.109 micromol.min(-1).g(-1) S9 protein or 8.62 +/- 2.02mol.min(-1).mol(-1) CYP, but was undetectable in parental CHL cell. CONCLUSION: The cDNA of human CYP2C9 was successfully cloned and a cell line of CHL- CYP2C9, efficiently expressing the protein of CYP2C9, was established.
文摘Bicoid is one of the important Drosophila maternal genes involved in the control of embryo polarity and larvae segmentation. To clone and characterize the rice bicoid-related genes, one cDNA clone, Rb24 (EMBL accession number: AJ2771380), was isolated by screening of rice unmature seed cDNA library. Sequence analysis indicates that Rb24 contains a putative amino acid sequence, which is homologous to unique 8 amino acids sequence within Drosophila bicoid homeodomain (50% identity, 75% similarity) and involves a lys-9 in putative helix 3. Northern blot analysis of rice RNA has shown that this sequence is expressed in a tissue-specific manner. The transcript was detected strongly in young panicles, but less in young leaves and roots. This results are further confirmed with paraffin section in situ hybridization. The signal is intensive in rice globular embryo and located at the apical tip of the embryo, then, along with the development of embryo, the signal is getting reduced and transfers into both sides of embryo. The existence of bicoid-related sequence in rice embryo and the similarity of polar distribution of bicoid and Rb24 mRNA in early embryo development may implicates a conserved maternal regulation mechanism of body axis presents in Drosophila and in rice.
基金supported by grants from the Ministry of Education and Science of Ukraine(0116U003593)grant from cieA3(Campus de Excelencia Internacional Agroalimentario)-UCO,Spain
文摘Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules. Despite extensive research on nsLTP in different plant species, mostly angiosperm, and the great diversity of physiological processes in which they seem to be involved, their exact functions still remain unclear. Also, very limited experimental data are available on nsLTP in gymnosperm. In this study, we report for the first time on the molecular cloning of nsLTP, from Pinus sylvestris L.(PsLTP1, GenBank accession JN980402.1) and the expression pattern of PsLTP1 during ontogenesis and in response to environmental stress conditions. Total RNA from roots of 7-day old pine seedlings was used to isolate the cDNA clone, corresponding to Scots pine lipid transfer protein. The open reading frame of PsLTP1 consists of 372 bp encoding a protein of 123 amino acids. Amino acid sequence alignment revealed that mature PsLTP1 shares high level of similarity with nsLTP from other conifers and with well-studied nsLTPs from angiosperms. The PsLTP1 contains a 27-amino-acid N-terminal signal sequence and presents all the features of a plant nsLTP. Amino acid comparison analysis and 3D structure prediction showed that PsLTP1 is a type 1 nsLTP. The results of the expression analysis of Scots pine PsLTP1 gene revealed that its transcripts accumulate in actively growing tissues. Furthermore, transcription of PsLTP1 was upregulated in response to cold and salt treatments, and downregulated during acidic, osmotic and water stresses.
基金the Natural Scientific Foundation of China (NSFC3962526)National High-Technology Project-863 (102-10-01-04)
文摘AIM: To develop and optimize cDNA representational difference analysis (cDNA RDA) method and to identify and clone garlic up-regulated genes in human gastric cancer (HGC) cells. METHODS: We performed cDNA RDA method by using abundant double-stranded cDNA messages provided by two self-constructed cDNA libraries (Allitridi-treated and paternal HGC cell line BGC823 cells cDNA libraries respectively). Bam H I and Xho I restriction sites harbored in the library vector were used to select representations. Northern and Slot blots analyses were employed to identify the obtained difference products. RESULTS: Fragments released from the cDNA library vector after restriction endonuclease digestion acted as good marker indicating the appropriate digestion degree for library DNA. Two novel expressed sequence tags (ESTs) and a recombinant gene were obtained. Slot blots result showed a 8-fold increase of glia-derived nexin/protease nexin 1 (GDN/PN1) gene expression level and 4-fold increase of hepatitis B virus x-interacting protein (XIP) mRNA level in BGC823 cells after Allitridi treatment for 72h. CONCLUSION: Elevated levels of GDN/PN1 and XIP mRNAs induced by Allitridi provide valuable molecular evidence for elucidating the garlic's efficacies against neurodegenerative and inflammatory diseases. Isolation of a recombinant gene and two novel ESTs further show cDNA RDA based on cDNA libraries to be a powerful method with high specificity and reproducibility in cloning differentially expressed genes.
基金Supported by the National Natural Science Foundation of China (Grant No. 30270896)
文摘Chemosensory proteins (CSPs) are ubiquitous soluble small proteins isolated from sensory organs thought to be involved in chemical communication. Here we report the first cDNA of CSPs, called Ac-ASP3, cloned and characterized from antennas of adult worker bees in Chinese honeybee, Apis cerana cerana. The Ac-ASP3 cDNA comprises 2 exons, with an ORF of 393-bp encoding 130 aa. Protein signature analyses show that the protein consists of four conserved cysteines and a signal peptide with 19 aa in the N-terminal sequence. The deduced protein sequence shares high homology with Am-ASP3 of Apis mellifera and low similarity with other species of insects. Immunocytochemical localization shows that Ac-ASP3 is only specifically expressed on the antenna contact chemosensilla such as sensilla trichodea B and sensilla basiconica, whereas Ac-ASP3 is scarcely expressed on olfactory chemosensilla such as sensilla placodea. Real-time PCR of Ac-ASP3 transcripts shows that Ac-ASP3 is highly expressed on wings and legs, but expression is lower on antenna. Temporal expression patterns suggest that Ac-ASP3 is expressed during the period of pupa and adults from 1-d to 6-d stages when bees act as house bees, cleaning the comb and taking care of the queen and larvae in comb. The above evidence suggests that Ac-ASP3 is unique in species and is generally not involved in olfaction during searching for honey and pollen. Rather, the protein seems to function in recognition of chemosensory substances on bees' cuticle and mechanical movement of antenna.
文摘The large protein of the hepatitis B virus (HBV) envelope was subdivided intothe preSl region with 108-115 amino acids, the preS2 region with 55 amino acidsand the S region with 226 amino acids. It was believed that the preSl region cotainedthe hepatocyte attachment sites of the virus as well as multiple highly immunogenicT-cell and B-cell epitopes. The antibody response to the preSl peptide was also oneof the most important serological markers of HBV infection.