RESUMEN
Genomic imprinting often results in parent-of-origin specific differential expression of maternally and paternally inherited alleles and plays an essential role in mammalian development and growth. Mammalian genomic imprinting has primarily been studied in mice and humans, with only limited information available for pigs. To systematically characterize this phenomenon and evaluate imprinting status between different species, we investigated imprinted genes on a genome-wide scale in pig brain tissues. Specifically, we performed bioinformatics analysis of high-throughput sequencing results from parental genomes and offspring transcriptomes of hybrid crosses between Duroc and Diannan small-ear pigs. We identified 11 paternally and five maternally expressed imprinted genes in pigs with highly stringent selection criteria. Additionally, we found that the KCNQ1 and IGF2R genes, which are related to development, displayed a different imprinting status in pigs compared with that in mice and humans. This comprehensive research should help improve our knowledge on genomic imprinting in pigs and highlight the potential use of imprinted genes in the pig breeding field.
Asunto(s)
Impresión Genómica , Mamíferos/genética , Porcinos/genética , Alelos , Animales , Estudio de Asociación del Genoma Completo , Especificidad de la EspecieRESUMEN
AIM: To prepare and characterize the monoclonal antibody against human GCRG213. METHODS: The HIS-GCRG213 fusion protein was expressed in E.coli. Mice were immunized with the purified HIS-GCRG213 protein. Hybridoma cell lines secreting monoclonal antibodies against GCRG213 were screened by regular cell fusion and subcloning approach. The titer and specificity of the antibody was characterized by ELISA and Western blot, respectively. The expression of GCRG213 was determined using immunohistochemistry technique on paraffin-embedded tissue sections from normal gastric mucosal tissues and advanced gastric cancer. RESULTS: The HIS-GCRG213 fusion protein with relative molecular mass of 20 800 was over expressed in E.coli. Two hybridoma cell lines which secreted monoclonal antibody specifically against human GCRG213 fusion protein were successfully obtained. The ascite titers of this monoclonal antibody reached 1:10(6);. Western blot analysis showed that the monoclonal antibody could bind to the recombinant HIS-GCRG213 protein specifically.The immunohistochemistry showed that GCRG213 were expressed higher in gastric cancer tissues than in normal ones. CONCLUSION: The monoclonal antibody against human GCRG213 with high titer and specificity has been successfully prepared, which could be utilized as a useful reagent for further studying the biological function of the GCRG213.
Asunto(s)
Anticuerpos Monoclonales/biosíntesis , Anticuerpos Monoclonales/inmunología , Hormonas Peptídicas/inmunología , Animales , Especificidad de Anticuerpos/inmunología , Escherichia coli/genética , Escherichia coli/metabolismo , Vectores Genéticos/genética , Humanos , Hibridomas/metabolismo , Ratones , Ratones Endogámicos BALB C , Hormonas Peptídicas/genética , Hormonas Peptídicas/aislamiento & purificación , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/aislamiento & purificación , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/metabolismoRESUMEN
AIM: To prepare the polyclonal antibody against gastric cancer-related protein GCRG224. METHODS: The thioredoxin/GCRG224 fusion protein was expressed in E.coli. The polyclonal antibody against GCRG224 was obtained by immunizing a rabbit with the purified GCRG224 protein. The titer and specificity of the antibody were determined by ELISA and Western blot, respectively. The expression of GCRG224 in paraffin-embedded tissue sections from normal gastric mucosal tissues and advanced gastric cancer was determined using immunohistochemistry technique. RESULTS: The thioredoxin/GCRG224 fusion protein with relative molecular mass of 16.8 kDa was over-expressed in E.coli. The purity of the expressed product directly purified from a denaturing polyacrylamide gel was about 100%. The polyclonal antibody against GCRG224 was prepared. ELISA detection proved the titer of antiserum against GCRG224 was about 1:256,000. Western blot analysis showed that the antiserum could bind to the expressed fusion protein specifically. GCRG224 was found to have higher expression in gastric cancer tissues than in normal ones by immunohistochemistry. CONCLUSION: The successful preparation of the polyclonal antibody against GCRG224 lays a foundation for further study of the biological function and the possible role of GCRG224 in the development of gastric carcinoma.
Asunto(s)
Anticuerpos/análisis , Anticuerpos/inmunología , Proteínas de Neoplasias/genética , Neoplasias Gástricas/genética , Animales , Anticuerpos/aislamiento & purificación , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Proteínas de Neoplasias/inmunología , Conejos , Neoplasias Gástricas/química , Neoplasias Gástricas/inmunologíaRESUMEN
AIM: To prepare the rabbit antibody against gastric cancer-related protein GCRG213. METHODS: The thioredoxin/GCRG213 fusion protein was expressed in E. coli. The rabbit antibody against GCRG213 was obtained by immunizing a rabbit with the purified GCRG213 protein. The titer and specificity of the antibody was determined by ELISA and Western-blot, respectively. RESULTS: The thioredoxin/GCRG213 fusion protein with relative molecular mass (Mr) of 29,400 was overexpressed in E. coli. The purity of expressed product directly purified from a denaturing polyacrylamide gel was about 100%. The rabbit antibody against GCRG213 was obtained. The ELISA titer of antiserum against GCRG213 was about 1:256,000. Western blot analysis showed that the antiserum could bind to the expressed fusion protein specifically. CONCLUSION: The rabbit antibody against GCRG213 has been successfully prepared, which lays the foundation for further studying the biological function and the possible role of the GCRG213 in the development of gastric carcinoma.