AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis ...AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR. RESULTS: HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion. CONCLUSION: HCV may exist and remain functional in a cultured cell line for a long period.展开更多
AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expr...AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expressed cDNAs in EB virus transformed human B cell.Then the isolated genes were cloned and sequenced and identifed by RT PCR.Nucleic acid homology searches were performed using the BLAST program.RESULTS:By this technique,4 differentiately expressed gene cDNA fragments of EB virus transformed human B cell were obtained.CONCLUSION:SSH is an effective method to isolate differentiately expressed genes.展开更多
基金The paper was support by a grant from the Ministry Youth Research of China,No.98-1-269
文摘AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR. RESULTS: HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion. CONCLUSION: HCV may exist and remain functional in a cultured cell line for a long period.
文摘AIM:To clone and identify genes differentially expressed in the EB virus transformed human B cell.METHODS:Suppression subtractive hybridization was used to construct the library which contains the differentiately expressed cDNAs in EB virus transformed human B cell.Then the isolated genes were cloned and sequenced and identifed by RT PCR.Nucleic acid homology searches were performed using the BLAST program.RESULTS:By this technique,4 differentiately expressed gene cDNA fragments of EB virus transformed human B cell were obtained.CONCLUSION:SSH is an effective method to isolate differentiately expressed genes.
