目的:CFI基因编码的补体因子I (FI)是补体系统调节过程中重要的丝氨酸蛋白酶,其功能缺陷与多种疾病相关,然而许多突变的致病机制并未完全阐明。本实验旨在通过迷你基因实验验证CFI基因突变对mRNA剪接的影响,为解释致病机制、发现治疗靶...目的:CFI基因编码的补体因子I (FI)是补体系统调节过程中重要的丝氨酸蛋白酶,其功能缺陷与多种疾病相关,然而许多突变的致病机制并未完全阐明。本实验旨在通过迷你基因实验验证CFI基因突变对mRNA剪接的影响,为解释致病机制、发现治疗靶点提供新思路。方法:本研究首先通过生物信息学软件,从人类基因数据库中筛选出目标突变,并将包含该突变位点的基因片段导入载体中。随后将构建好的含有野生型和突变型的质粒分别转染至人胚肾细胞(HEK293T)中,并利用RT-PCR和测序技术分析其mRNA产物的变化。结果:与野生型5号外显子相比,c.772G>A突变导致外显子完全跳变,产生了异常剪接产物。这一异常剪接可能导致产生的FI蛋白功能域缺陷,从而影响其蛋白酶活性或与其他补体因子的结合。结论:本研究成功构建了CFI基因的迷你基因实验体系,证实了c.772G>A突变可导致mRNA剪接异常。这为深入理解CFI基因突变的致病机制提供了重要的实验依据,也为aHUS疾病的基因诊断和治疗提供了新的靶点。 Objective: Complement factor I (FI), encoded by the CFI gene, is an important serine protease regulating the complement system, and its functional defects have been associated with a wide range of diseases. However, the pathogenic mechanisms of many mutations have not been fully elucidated. This experiment aims to verify the effect of CFI gene mutation on mRNA splicing through minigene experiments, which will provide new ideas to explain the pathogenic mechanism and discover therapeutic targets. Methods: In this study, the target mutation was first screened out from the human gene database by bioinformatics software, and the gene fragment containing the mutation site was introduced into the vector. Subsequently, the constructed plasmids containing wild-type and mutant were transfected into human embryonic kidney cells (HEK293T) respectively, and the changes in their mRNA products were analyzed by RT-PCR and sequencing. Results: Compared with wild-type exon 5, the c.772G>A mutation resulted in a complete exon skipping, producing an aberrant splicing product. This aberrant splicing may lead to defects in the functional domains of the resulting FI proteins, thus affecting their protease activity or binding to other complement factors. Conclusion: In this study, we successfully constructed a minigene experimental system for the CFI gene and confirmed that the c.772G>A mutation could lead to abnormal mRNA splicing. This provides an essential experimental basis for the in-depth understanding of the pathogenic mechanism of CFI gene mutation and also provides a new target for the genetic diagnosis and treatment of aHUS disease.展开更多
On the occasion of UNITECR’07 to be held on Sept.18-21, 2007, Ceramic Forum International (CFI) will publish a special issue for refractories. Following are abstracts form the issue for a first look for our readers.
重点研究了LTE系统中存在承载PDCCH(physical downlink control channel,物理下行控制信道)传输的信息冗余和PDCCH盲检效率较低的问题,提出了一种改进的计算CFI值的方法。该方法按照速率匹配值的降序为各个PDCCH分配1/2/4/8个CCE(contro...重点研究了LTE系统中存在承载PDCCH(physical downlink control channel,物理下行控制信道)传输的信息冗余和PDCCH盲检效率较低的问题,提出了一种改进的计算CFI值的方法。该方法按照速率匹配值的降序为各个PDCCH分配1/2/4/8个CCE(control channel element,控制信道元素),再复用各个PDCCH。仿真结果表明该方法能够有效地消除冗余信息,减少PDCCH盲检的平均次数。展开更多
2004年7月25日至8月8日,笔者来到风光绮丽的美国纽约州临近布法罗市的阿姆赫斯特,对坐落在那里探索中心(Center for Inquiry,简称CFI)进行了为期两周的访问与学习.笔者访问CFI整个机构,参观了图书馆,访问了普洛米休斯出版社,与著名... 2004年7月25日至8月8日,笔者来到风光绮丽的美国纽约州临近布法罗市的阿姆赫斯特,对坐落在那里探索中心(Center for Inquiry,简称CFI)进行了为期两周的访问与学习.笔者访问CFI整个机构,参观了图书馆,访问了普洛米休斯出版社,与著名学者进行了两次学术交流,完成了一个介绍中国探索小组工作以及CFI如何帮助中国学者推动批评性思考在中国进行的小项目.……展开更多
文摘目的:CFI基因编码的补体因子I (FI)是补体系统调节过程中重要的丝氨酸蛋白酶,其功能缺陷与多种疾病相关,然而许多突变的致病机制并未完全阐明。本实验旨在通过迷你基因实验验证CFI基因突变对mRNA剪接的影响,为解释致病机制、发现治疗靶点提供新思路。方法:本研究首先通过生物信息学软件,从人类基因数据库中筛选出目标突变,并将包含该突变位点的基因片段导入载体中。随后将构建好的含有野生型和突变型的质粒分别转染至人胚肾细胞(HEK293T)中,并利用RT-PCR和测序技术分析其mRNA产物的变化。结果:与野生型5号外显子相比,c.772G>A突变导致外显子完全跳变,产生了异常剪接产物。这一异常剪接可能导致产生的FI蛋白功能域缺陷,从而影响其蛋白酶活性或与其他补体因子的结合。结论:本研究成功构建了CFI基因的迷你基因实验体系,证实了c.772G>A突变可导致mRNA剪接异常。这为深入理解CFI基因突变的致病机制提供了重要的实验依据,也为aHUS疾病的基因诊断和治疗提供了新的靶点。 Objective: Complement factor I (FI), encoded by the CFI gene, is an important serine protease regulating the complement system, and its functional defects have been associated with a wide range of diseases. However, the pathogenic mechanisms of many mutations have not been fully elucidated. This experiment aims to verify the effect of CFI gene mutation on mRNA splicing through minigene experiments, which will provide new ideas to explain the pathogenic mechanism and discover therapeutic targets. Methods: In this study, the target mutation was first screened out from the human gene database by bioinformatics software, and the gene fragment containing the mutation site was introduced into the vector. Subsequently, the constructed plasmids containing wild-type and mutant were transfected into human embryonic kidney cells (HEK293T) respectively, and the changes in their mRNA products were analyzed by RT-PCR and sequencing. Results: Compared with wild-type exon 5, the c.772G>A mutation resulted in a complete exon skipping, producing an aberrant splicing product. This aberrant splicing may lead to defects in the functional domains of the resulting FI proteins, thus affecting their protease activity or binding to other complement factors. Conclusion: In this study, we successfully constructed a minigene experimental system for the CFI gene and confirmed that the c.772G>A mutation could lead to abnormal mRNA splicing. This provides an essential experimental basis for the in-depth understanding of the pathogenic mechanism of CFI gene mutation and also provides a new target for the genetic diagnosis and treatment of aHUS disease.
基金supported by the Natural Science Basic Research Plan in Shaanxi Province of China(No.2014JQ1015)the Funding Project on the Key Supporting Subject of Mathematics in Shaanxi Province of China(No.14SXZD003)+1 种基金the Specialized Research Fund for the Doctoral Program of Higher Education(No.20110202110002)the Science and Technology Planning Project of Weinan City(No.2013KYJ-1)
文摘On the occasion of UNITECR’07 to be held on Sept.18-21, 2007, Ceramic Forum International (CFI) will publish a special issue for refractories. Following are abstracts form the issue for a first look for our readers.
文摘 2004年7月25日至8月8日,笔者来到风光绮丽的美国纽约州临近布法罗市的阿姆赫斯特,对坐落在那里探索中心(Center for Inquiry,简称CFI)进行了为期两周的访问与学习.笔者访问CFI整个机构,参观了图书馆,访问了普洛米休斯出版社,与著名学者进行了两次学术交流,完成了一个介绍中国探索小组工作以及CFI如何帮助中国学者推动批评性思考在中国进行的小项目.……
