To detect mutations of the aquaporin 2 gene(AQP2) and the arginine vasopressin V2 receptor gene(AVPR2) of Chinese congenital nephrogenic diabetes insipidus, and to establish the foundation for further studying the...To detect mutations of the aquaporin 2 gene(AQP2) and the arginine vasopressin V2 receptor gene(AVPR2) of Chinese congenital nephrogenic diabetes insipidus, and to establish the foundation for further studying the emergence mechanism of the disease and clinical diagnosis, all the exons and part of introns of AQP2 and AVPR2 genes were amplified with intronic primers, using genomic DNA extracted from three patients with congenital nephrogenic diabetes insipidus and two mothers as template, PCR product was ligated into a T-vector and then sequenced. The result was compared with the database sequence to identify the mutable sites via a BLAST search, the incidence of every mutation was analyzed, and the putative transcription factor binding sites that maybe disturbed were analyzed by MAPPER. Mutation g.1394A〉G in exon 3 of AVPR2 was detected in all the subjects, g.861C〉T(S167L) in exon 2 of AVPR2 and IVS1+3G〉A in intron of AQP2 were detected, respectively, in two patients, and c.836A〉C in 3′ untranslated region of AQP2 was detected in two patients and one mother. Four mutations were identified. g.1394A〉G of AVPR2 and c.836A〉C of AQP2 have high incidence in patients with nephrogenic diabetes insipidus. Detection on the two sites may become auxiliary diagnosis index of congenital nephrogenic diabetes insipidus.展开更多
The root of Panax ginseng plant undergoes a specific developmental process to become a biosynthesis and accumulation tissue for ginsenosides. To identify and analyze genes involved in the biosynthesis of ginsenoside, ...The root of Panax ginseng plant undergoes a specific developmental process to become a biosynthesis and accumulation tissue for ginsenosides. To identify and analyze genes involved in the biosynthesis of ginsenoside, we constructed and characterized a full-length cDNA library for 6-year-old North American ginseng. The titer of primary cDNA library is 1.2 × 10^6 pfu/mL, the titer of amplified library is 2. 6 × 10^10 pfu/mL and the rate of recombinant is above 86%. The insert size ranges from 0. 3 to 2.0 kb. Sequencing results show that 18 of 58 genes are high homologous to the genes (GBRS, GBR3 and GBR1 ) known in GenBank, which are involved in biosynthesis of ginsenoside in North American ginseng plant; 16 of 58 genes are novel genes. The full-length cDNA library of North American ginseng root tissues is essential for the cloning of genes known and it is also an initial key for the screening and cloning of new genes.展开更多
Testis-specific protease 50 (TSP50) is a testis-specific oncogene, which is abnormally activated in most tested patients with breast cancer. This property makes it an attractive molecular marker and a promising targ...Testis-specific protease 50 (TSP50) is a testis-specific oncogene, which is abnormally activated in most tested patients with breast cancer. This property makes it an attractive molecular marker and a promising target for the diagnosis and therapy of breast cancer. In order to obtain the protective and specific polyclonal antibodies for further research, TSPS0 cDNA was amplified by RT-PCR from normal human testicular tissue, and inserted into eukaryotic expression vector PeDNA3.1. Rabbit anti-TSPS0 polyclonal antibodies were prepared by means of intramuscular injection of peDNA3.1-TSPS0 into the rabbits. Titem of the anti-sera were measured by ELISA and Western blotting with the E. coli cell lysate containing the induced GST-TSPS0 fusion protein as an antigen. In addition, we examined the expression of TSPS0 in both breast cancer cell line MCF-7 and breast cancer tissue by immunofluorescent and immunohistochemistry analysis.展开更多
Testis-specific protease 50(TSP50) has been identified as a testis-specific protein that is expressed abnormally in most human breast cancer samples,which makes it an attractive molecular marker and a potential targ...Testis-specific protease 50(TSP50) has been identified as a testis-specific protein that is expressed abnormally in most human breast cancer samples,which makes it an attractive molecular marker and a potential target for diagnosis and therapy.In the present study,we prepared a panel of monoclonal antibodies(mAbs) with high specificity and sensitivity against TSP50 by hybridoma method and characterized them by ELISA,Western blot,immunofluroescence and immunohistochemical analyses.The results show that all of the 9 different clones can specifically bind to TSP50.The mAbs against TSP50 we generated could be good tools for both basic and clinical studies.展开更多
基金the National High Technology Research and Development Program of China(No.2004AA2Z3782)Key Grant of Jilin Science & Technology Committee(No.20060904)+2 种基金Program for New Century Excellent Talents in Universities of China (No.NCET-06-0320)Foundation of Fostering Science and Technology Innovation Program of Northest Normal University (No.NENU-STB07008)Analysis and Testing Foundation of Northest Normal University.
文摘To detect mutations of the aquaporin 2 gene(AQP2) and the arginine vasopressin V2 receptor gene(AVPR2) of Chinese congenital nephrogenic diabetes insipidus, and to establish the foundation for further studying the emergence mechanism of the disease and clinical diagnosis, all the exons and part of introns of AQP2 and AVPR2 genes were amplified with intronic primers, using genomic DNA extracted from three patients with congenital nephrogenic diabetes insipidus and two mothers as template, PCR product was ligated into a T-vector and then sequenced. The result was compared with the database sequence to identify the mutable sites via a BLAST search, the incidence of every mutation was analyzed, and the putative transcription factor binding sites that maybe disturbed were analyzed by MAPPER. Mutation g.1394A〉G in exon 3 of AVPR2 was detected in all the subjects, g.861C〉T(S167L) in exon 2 of AVPR2 and IVS1+3G〉A in intron of AQP2 were detected, respectively, in two patients, and c.836A〉C in 3′ untranslated region of AQP2 was detected in two patients and one mother. Four mutations were identified. g.1394A〉G of AVPR2 and c.836A〉C of AQP2 have high incidence in patients with nephrogenic diabetes insipidus. Detection on the two sites may become auxiliary diagnosis index of congenital nephrogenic diabetes insipidus.
基金Supported by the Science Fund for Social Development Program(Modern Chinese Medicine) of Jilin Province(No20011109) Analysis and Testing Foundation of Northeast Normal University
文摘The root of Panax ginseng plant undergoes a specific developmental process to become a biosynthesis and accumulation tissue for ginsenosides. To identify and analyze genes involved in the biosynthesis of ginsenoside, we constructed and characterized a full-length cDNA library for 6-year-old North American ginseng. The titer of primary cDNA library is 1.2 × 10^6 pfu/mL, the titer of amplified library is 2. 6 × 10^10 pfu/mL and the rate of recombinant is above 86%. The insert size ranges from 0. 3 to 2.0 kb. Sequencing results show that 18 of 58 genes are high homologous to the genes (GBRS, GBR3 and GBR1 ) known in GenBank, which are involved in biosynthesis of ginsenoside in North American ginseng plant; 16 of 58 genes are novel genes. The full-length cDNA library of North American ginseng root tissues is essential for the cloning of genes known and it is also an initial key for the screening and cloning of new genes.
文摘Testis-specific protease 50 (TSP50) is a testis-specific oncogene, which is abnormally activated in most tested patients with breast cancer. This property makes it an attractive molecular marker and a promising target for the diagnosis and therapy of breast cancer. In order to obtain the protective and specific polyclonal antibodies for further research, TSPS0 cDNA was amplified by RT-PCR from normal human testicular tissue, and inserted into eukaryotic expression vector PeDNA3.1. Rabbit anti-TSPS0 polyclonal antibodies were prepared by means of intramuscular injection of peDNA3.1-TSPS0 into the rabbits. Titem of the anti-sera were measured by ELISA and Western blotting with the E. coli cell lysate containing the induced GST-TSPS0 fusion protein as an antigen. In addition, we examined the expression of TSPS0 in both breast cancer cell line MCF-7 and breast cancer tissue by immunofluorescent and immunohistochemistry analysis.
基金Supported by the National Natural Science Foundation of China(No.30672068)the Distinguished Young Scholars Fund of Jilin Province,China(No.20050114)+1 种基金the Key Grant of Jilin Province Science & Technology Committee,China(Nos.20060923-01,200805131)the Key Grant of Changchun City Science & Technology Committee,China(No.06GG147)
文摘Testis-specific protease 50(TSP50) has been identified as a testis-specific protein that is expressed abnormally in most human breast cancer samples,which makes it an attractive molecular marker and a potential target for diagnosis and therapy.In the present study,we prepared a panel of monoclonal antibodies(mAbs) with high specificity and sensitivity against TSP50 by hybridoma method and characterized them by ELISA,Western blot,immunofluroescence and immunohistochemical analyses.The results show that all of the 9 different clones can specifically bind to TSP50.The mAbs against TSP50 we generated could be good tools for both basic and clinical studies.