目的:探讨溴化乙锭(EB)诱导法建立人线粒体DNA(mtDNA)缺失宫颈癌Hela S3细胞,以及再转入线粒体构建融合细胞的可行性,并对转线粒体细胞进行初步分析。方法:采用低剂量(100 ng/m L)EB诱导法建立mtDNA缺失的ρ0 Hela S3细胞,通过普通PCR...目的:探讨溴化乙锭(EB)诱导法建立人线粒体DNA(mtDNA)缺失宫颈癌Hela S3细胞,以及再转入线粒体构建融合细胞的可行性,并对转线粒体细胞进行初步分析。方法:采用低剂量(100 ng/m L)EB诱导法建立mtDNA缺失的ρ0 Hela S3细胞,通过普通PCR、荧光定量PCR(qPCR)进行mtDNA缺失鉴定。采用聚乙二醇(PEG)介导细胞融合法,以正常人血小板为mtDNA供体转入ρ0 Hela S3细胞,构建转线粒体细胞(transmitochondrial cybrids),并通过PCR、qPCR和透射电镜观察进行初步分析和鉴定。结果:普通PCR和qPCR结果证实EB诱导法能够成功建立ρ0 Hela S3细胞,同时结合透射电镜证明转线粒体细胞能够恢复线粒体正常结构。结论:采用EB诱导法可成功建立ρ0 Hela S3细胞,且通过细胞融合构建的转线粒体细胞能够恢复mtDNA复制和正常线粒体结构,为研究mtDNA突变与线粒体功能异常相关疾病的关系提供可靠的实验基础。展开更多
The authors sequenced POLG1,C10ORF2,and ANT1 in 38 sporadic progressive external ophthalmoplegia patients withmultiple mitochondrial DNA(mtDNA)deletions.Cau-sative mutations were identified in approximately 10% of cas...The authors sequenced POLG1,C10ORF2,and ANT1 in 38 sporadic progressive external ophthalmoplegia patients withmultiple mitochondrial DNA(mtDNA)deletions.Cau-sative mutations were identified in approximately 10% of cases,with two unrelated individuals harboring a novel premature stop codon mutation(1356T > G).None had a mutation in C10ORF2 or ANT1.In the majority of patients,the primary nuclear genetic defect is likely to affect other unknown genes important for mtDNA maintenance.展开更多
The authors sequenced POLG1, C10ORF2, and ANT1 in 38 sporadic progressive external ophthalmoplegia patients with multiple mitochondrial DNA (mtDNA) deletions. Causative mutations were identified in approximately 10%of...The authors sequenced POLG1, C10ORF2, and ANT1 in 38 sporadic progressive external ophthalmoplegia patients with multiple mitochondrial DNA (mtDNA) deletions. Causative mutations were identified in approximately 10%of cases, with two unrelated individuals harboring a novel premature stop codon mutation (1356T > G). None had a mutation in C10ORF2 or ANT1. In the majority of patients, the primary nuclear genetic defect is likely to affect other unknown genes important for mtDNA maintenance.展开更多
文摘目的:探讨溴化乙锭(EB)诱导法建立人线粒体DNA(mtDNA)缺失宫颈癌Hela S3细胞,以及再转入线粒体构建融合细胞的可行性,并对转线粒体细胞进行初步分析。方法:采用低剂量(100 ng/m L)EB诱导法建立mtDNA缺失的ρ0 Hela S3细胞,通过普通PCR、荧光定量PCR(qPCR)进行mtDNA缺失鉴定。采用聚乙二醇(PEG)介导细胞融合法,以正常人血小板为mtDNA供体转入ρ0 Hela S3细胞,构建转线粒体细胞(transmitochondrial cybrids),并通过PCR、qPCR和透射电镜观察进行初步分析和鉴定。结果:普通PCR和qPCR结果证实EB诱导法能够成功建立ρ0 Hela S3细胞,同时结合透射电镜证明转线粒体细胞能够恢复线粒体正常结构。结论:采用EB诱导法可成功建立ρ0 Hela S3细胞,且通过细胞融合构建的转线粒体细胞能够恢复mtDNA复制和正常线粒体结构,为研究mtDNA突变与线粒体功能异常相关疾病的关系提供可靠的实验基础。
文摘The authors sequenced POLG1,C10ORF2,and ANT1 in 38 sporadic progressive external ophthalmoplegia patients withmultiple mitochondrial DNA(mtDNA)deletions.Cau-sative mutations were identified in approximately 10% of cases,with two unrelated individuals harboring a novel premature stop codon mutation(1356T > G).None had a mutation in C10ORF2 or ANT1.In the majority of patients,the primary nuclear genetic defect is likely to affect other unknown genes important for mtDNA maintenance.
文摘The authors sequenced POLG1, C10ORF2, and ANT1 in 38 sporadic progressive external ophthalmoplegia patients with multiple mitochondrial DNA (mtDNA) deletions. Causative mutations were identified in approximately 10%of cases, with two unrelated individuals harboring a novel premature stop codon mutation (1356T > G). None had a mutation in C10ORF2 or ANT1. In the majority of patients, the primary nuclear genetic defect is likely to affect other unknown genes important for mtDNA maintenance.
