[目的 ]扩增和克隆猪带绦虫囊尾蚴 Ag B基因的 c DNA编码区。 [方法 ]提取猪囊尾蚴总 RNA中 ,利用 RT- PCR技术扩增出 Ag B基因 c DNA编码区 ,然后将其克隆到载体 p UC118中进行序列分析。 [结果 ]PCR反应产物为单一条带 ,大小为 2 .6 k...[目的 ]扩增和克隆猪带绦虫囊尾蚴 Ag B基因的 c DNA编码区。 [方法 ]提取猪囊尾蚴总 RNA中 ,利用 RT- PCR技术扩增出 Ag B基因 c DNA编码区 ,然后将其克隆到载体 p UC118中进行序列分析。 [结果 ]PCR反应产物为单一条带 ,大小为 2 .6 kb。测序结果与澳大利亚株猪囊尾蚴 Ag B序列有 99.8%的同源性 ,二者的氨基酸序列有 99.3%的同源性。 [结论 ]成功地克隆了猪囊尾蚴 Ag B基因 c DNA编码区。展开更多
UDP-glucose pyrophosphorylase is an important enzyme concerned with carbohydrate metabolism in plants. Cloning of UGPase is a premise for further study on molecular level, and it is also crucial for study of carbohydr...UDP-glucose pyrophosphorylase is an important enzyme concerned with carbohydrate metabolism in plants. Cloning of UGPase is a premise for further study on molecular level, and it is also crucial for study of carbohydrate metabolism. UGPase cDNA sequence as a template, designed primer, then 3＇-untranslate region （3＇ UTR） of UGPase were amplified by 3＇-rapid amplification of cDNA ends （3＇-RACE）. The results suggested the 3＇ UTR were 243 bp, contained AATAA sequence and Poly（A）.展开更多
Bactericidal/permeability-increasing protein （BPI） and LPS-binding protein （LBP） play an important role in host defence. Current evidence shows that BPI/LBP may be widely existed in different cells and tissue type...Bactericidal/permeability-increasing protein （BPI） and LPS-binding protein （LBP） play an important role in host defence. Current evidence shows that BPI/LBP may be widely existed in different cells and tissue types of animals. A full-length cDNA clone encoding a BPI/LBP homologue （dBPI）, 1757 bp in size, was characterized in venom gland of the hundred-pace snake Deinagkistrodon acutus. Its deduced amino acid sequence of 417 residues had 13.8% - 21.5% identity to BPI like 1 （BPIL1） and BPI like 3 （BPIL3） of other animals. Conserved cysteine residues which are involved in disulfide bond formation between the final strand of the N-terminal beta sheet and the long alpha helix of BPI are identified as Cys146-Cys183 of dBPI. Phylogenetic tree analysis showed that the BPI/LBP homologues formed five large clusters and dBPI was in a large cluster including BPIL1 and BPIL3. dBPI mRNA shows a tissue specific expression in venom gland. This is the first study to identify the cDNA encoding BPI/LBP homologues from reptiles [ Current Zoology 55 （5） ： 376 - 382, 2009].展开更多
Galectins, a family of β-galactoside-binding proteins, participate in both innate immunity and adaptive immunity. This study identified one novel galectin-related protein from half-smooth tongue sole Cynoglossus semi...Galectins, a family of β-galactoside-binding proteins, participate in both innate immunity and adaptive immunity. This study identified one novel galectin-related protein from half-smooth tongue sole Cynoglossus semilaevis, which was designated as CsGRP. The full-length cDNA of its encoding gene was 785 bp in length with a 528 bp open reading frame encoding a putative protein of 176 amino acids. The deduced CsGRP contained a putative 131-aa galactoside-binding lectin domain and 3 critical residues responsible for carbohydrate binding （R93, W109 and R114）. Genomic structural analysis revealed that CsGRP consisted of five exons and four intzons. CsGRP showed 68% similarity with Poecilia latipinna GRP and 67% similarity wih Stegastes partitus GRP. CsGRP showed the highest expression level in liver, although its expression was detected in all tested tissues. When challenged with Vibrio harveyi, the expression of CsGRP was significantly down-regulated in liver （P〈 0.05）. In addition, we found that in spleen and kidney of C. semilaevis, the CpG island of CsGRP showed significantly higher （P 〈 0.05） methylation level in disease-resistant family of C. semilaevis （DR-Cs） than in disease-susceptible ones （DS-Cs）. Our results suggested that CsGRP may play important roles in the immune response of C. semilaevis. Moreover, DNA methylation patterns provided valuable data for understanding the relationship between epigenetic regulation and immunity, which would assist the animal genetic research and improve the animal breeding in the future.展开更多
The total RNA was extracted from Microtus fortis liver tissue which before being infected and after being infected 10 d and 15 d by the Schistosoma japonicum cercariae. Using rattus norvegicus CD72 gene probes were us...The total RNA was extracted from Microtus fortis liver tissue which before being infected and after being infected 10 d and 15 d by the Schistosoma japonicum cercariae. Using rattus norvegicus CD72 gene probes were used to hybridize analysis of CD72 difference expression in the Microtus fortis liver tissues which were infected with Schistosoma japonicum before and after being infected. At the same time, the cDNA sequence and encoded amino acid sequence of the rattus norvegicus CD72 gene and CD72 protein structural domains were analyzed by using bioinformatics. The results showed that the CD72 expression levels in the liver tissue of Microtus fortis after being infected was significantly higher than before being infected. The rattus norvegicus CD72 cDNA sequence of a total length is 1479 bp and encode 364 amino acid residues and rattus norvegicus CD72 protein containing a CD72 superfamily domain.展开更多
Cyclin B is an important regulator of the cell cycle G2 to M phase transition. The silkworm genomic database shows that there are two Cyclin B genes in the silkworm (Bombyx mori), BmCyclin B and BmCyclin B3. Using sil...Cyclin B is an important regulator of the cell cycle G2 to M phase transition. The silkworm genomic database shows that there are two Cyclin B genes in the silkworm (Bombyx mori), BmCyclin B and BmCyclin B3. Using silkworm EST data, the cyclin B3 (EU074796) gene was cloned. Its complete cDNA was 1665 bp with an ORF of 1536 bp derived from seven exons and six introns. The BmCyclin B3 gene encodes 511 amino acids, and the predicted molecular weight is 57.8 kD with an isoelectric point of 9.18. The protein contains one protein damage box and two cyclin boxes. RNA interference-mediated reduction of BmCyclin B and BmCyclin B3 expression induced cell cycle arrest in G2 or M phase in BmN-SWU1 cells, thus inhibiting cell proliferation. These results suggest that BmCyclin B and BmCyclin B3 are necessary for completing the cell cycle in silkworm cells.展开更多
β-carotene ketolase and β-carotene hydroxylase encoded by bkt and bch, respectively, are key enzymes required for astaxanthin biosynthesis in Haematococcu pluvialis 34-1n. Two expression vectors containing cDNA sequ...β-carotene ketolase and β-carotene hydroxylase encoded by bkt and bch, respectively, are key enzymes required for astaxanthin biosynthesis in Haematococcu pluvialis 34-1n. Two expression vectors containing cDNA sequences of bkt and bch were constructed and co-transformed into cell-wall-deficient Chlamydomonas reinhardtii CC-849. Transgenic algae were screened on TAP agar plates containing 10 gg mL 1 Zeomycin. PCR-Southern analysis showed that bkt and bch were integrated into the genomes of C. reinhardtii. Transcripts of bkt and bch were further confirmed by RT-PCR-Southern analysis. Compared with the wild type, transgenic algae produced 29.04% and 30.27% more carotenoids and xanthophylls, respectively. Moreover, the transgenic algae could accumulate 34% more astaxanthin than wild type. These results indicate that foreign bkt and bch genes were successfully translated into β-carotene ketolase and β-carotene hydroxylase, which were responsible for catalyzing the biosynthesis of astaxanthin in transgenic algae.展开更多
文摘[目的 ]扩增和克隆猪带绦虫囊尾蚴 Ag B基因的 c DNA编码区。 [方法 ]提取猪囊尾蚴总 RNA中 ,利用 RT- PCR技术扩增出 Ag B基因 c DNA编码区 ,然后将其克隆到载体 p UC118中进行序列分析。 [结果 ]PCR反应产物为单一条带 ,大小为 2 .6 kb。测序结果与澳大利亚株猪囊尾蚴 Ag B序列有 99.8%的同源性 ,二者的氨基酸序列有 99.3%的同源性。 [结论 ]成功地克隆了猪囊尾蚴 Ag B基因 c DNA编码区。
文摘UDP-glucose pyrophosphorylase is an important enzyme concerned with carbohydrate metabolism in plants. Cloning of UGPase is a premise for further study on molecular level, and it is also crucial for study of carbohydrate metabolism. UGPase cDNA sequence as a template, designed primer, then 3＇-untranslate region （3＇ UTR） of UGPase were amplified by 3＇-rapid amplification of cDNA ends （3＇-RACE）. The results suggested the 3＇ UTR were 243 bp, contained AATAA sequence and Poly（A）.
基金funded by a grant from the local government of Zhejiang Province for the Specially Supported Discipline of Zoology
文摘Bactericidal/permeability-increasing protein （BPI） and LPS-binding protein （LBP） play an important role in host defence. Current evidence shows that BPI/LBP may be widely existed in different cells and tissue types of animals. A full-length cDNA clone encoding a BPI/LBP homologue （dBPI）, 1757 bp in size, was characterized in venom gland of the hundred-pace snake Deinagkistrodon acutus. Its deduced amino acid sequence of 417 residues had 13.8% - 21.5% identity to BPI like 1 （BPIL1） and BPI like 3 （BPIL3） of other animals. Conserved cysteine residues which are involved in disulfide bond formation between the final strand of the N-terminal beta sheet and the long alpha helix of BPI are identified as Cys146-Cys183 of dBPI. Phylogenetic tree analysis showed that the BPI/LBP homologues formed five large clusters and dBPI was in a large cluster including BPIL1 and BPIL3. dBPI mRNA shows a tissue specific expression in venom gland. This is the first study to identify the cDNA encoding BPI/LBP homologues from reptiles [ Current Zoology 55 （5） ： 376 - 382, 2009].
基金supported by the National Natural Science Foundation(No.31530078)the Central Public-Interest Scientific Institution Basal Research Fund,CAFS(Nos.2016HY-ZD0201,2017GH02)+4 种基金the China Agriculture Research System(No.CARS-47-G03)the Taishan Scholar Climbing Project Fund of Shandong,Chinathe China Postdoctoral Science Foundation(No.2015M582171)the Post-Doctoral Applied Research Project Fund of Qingdao CityApplied Basic Research Project of Qingdao City(No.16-5-1-52-jch)
文摘Galectins, a family of β-galactoside-binding proteins, participate in both innate immunity and adaptive immunity. This study identified one novel galectin-related protein from half-smooth tongue sole Cynoglossus semilaevis, which was designated as CsGRP. The full-length cDNA of its encoding gene was 785 bp in length with a 528 bp open reading frame encoding a putative protein of 176 amino acids. The deduced CsGRP contained a putative 131-aa galactoside-binding lectin domain and 3 critical residues responsible for carbohydrate binding （R93, W109 and R114）. Genomic structural analysis revealed that CsGRP consisted of five exons and four intzons. CsGRP showed 68% similarity with Poecilia latipinna GRP and 67% similarity wih Stegastes partitus GRP. CsGRP showed the highest expression level in liver, although its expression was detected in all tested tissues. When challenged with Vibrio harveyi, the expression of CsGRP was significantly down-regulated in liver （P〈 0.05）. In addition, we found that in spleen and kidney of C. semilaevis, the CpG island of CsGRP showed significantly higher （P 〈 0.05） methylation level in disease-resistant family of C. semilaevis （DR-Cs） than in disease-susceptible ones （DS-Cs）. Our results suggested that CsGRP may play important roles in the immune response of C. semilaevis. Moreover, DNA methylation patterns provided valuable data for understanding the relationship between epigenetic regulation and immunity, which would assist the animal genetic research and improve the animal breeding in the future.
文摘The total RNA was extracted from Microtus fortis liver tissue which before being infected and after being infected 10 d and 15 d by the Schistosoma japonicum cercariae. Using rattus norvegicus CD72 gene probes were used to hybridize analysis of CD72 difference expression in the Microtus fortis liver tissues which were infected with Schistosoma japonicum before and after being infected. At the same time, the cDNA sequence and encoded amino acid sequence of the rattus norvegicus CD72 gene and CD72 protein structural domains were analyzed by using bioinformatics. The results showed that the CD72 expression levels in the liver tissue of Microtus fortis after being infected was significantly higher than before being infected. The rattus norvegicus CD72 cDNA sequence of a total length is 1479 bp and encode 364 amino acid residues and rattus norvegicus CD72 protein containing a CD72 superfamily domain.
基金supported by the National Natural Science Foundation of China (31272505, 31172269)
文摘Cyclin B is an important regulator of the cell cycle G2 to M phase transition. The silkworm genomic database shows that there are two Cyclin B genes in the silkworm (Bombyx mori), BmCyclin B and BmCyclin B3. Using silkworm EST data, the cyclin B3 (EU074796) gene was cloned. Its complete cDNA was 1665 bp with an ORF of 1536 bp derived from seven exons and six introns. The BmCyclin B3 gene encodes 511 amino acids, and the predicted molecular weight is 57.8 kD with an isoelectric point of 9.18. The protein contains one protein damage box and two cyclin boxes. RNA interference-mediated reduction of BmCyclin B and BmCyclin B3 expression induced cell cycle arrest in G2 or M phase in BmN-SWU1 cells, thus inhibiting cell proliferation. These results suggest that BmCyclin B and BmCyclin B3 are necessary for completing the cell cycle in silkworm cells.
基金supported by the National Natural Science Foundation of China(41176106,31470389,31470431)Shenzhen Grant Plan for Science & Technology(CXB201104210005A,JCYJ20120613112512654,JSGG20130411160539208)Guangdong Enterprise Academician Workstation(2011A090700015)
文摘β-carotene ketolase and β-carotene hydroxylase encoded by bkt and bch, respectively, are key enzymes required for astaxanthin biosynthesis in Haematococcu pluvialis 34-1n. Two expression vectors containing cDNA sequences of bkt and bch were constructed and co-transformed into cell-wall-deficient Chlamydomonas reinhardtii CC-849. Transgenic algae were screened on TAP agar plates containing 10 gg mL 1 Zeomycin. PCR-Southern analysis showed that bkt and bch were integrated into the genomes of C. reinhardtii. Transcripts of bkt and bch were further confirmed by RT-PCR-Southern analysis. Compared with the wild type, transgenic algae produced 29.04% and 30.27% more carotenoids and xanthophylls, respectively. Moreover, the transgenic algae could accumulate 34% more astaxanthin than wild type. These results indicate that foreign bkt and bch genes were successfully translated into β-carotene ketolase and β-carotene hydroxylase, which were responsible for catalyzing the biosynthesis of astaxanthin in transgenic algae.
