Liver hepatocellular cancer(LIHC)is positioned as the third cancer with the highest mortalities worldwide,and high mortalities are associated with late diagnosis and recurrence.This study advances bioinformatics analy...Liver hepatocellular cancer(LIHC)is positioned as the third cancer with the highest mortalities worldwide,and high mortalities are associated with late diagnosis and recurrence.This study advances bioinformatics analysis of FAM3A expression in LIHC to evaluate its potential as a prognostic,diagnostic and therapeutic biomarker.Bioinformatics tools such as UALCAN,GEPIA2,KM plotter,TIMER2 and cBioPortal are employed to conduct analysis.Initially,the expression analysis revealed up-regulation of FAM3A in LIHC based on various variables.Further,the study observed that FAM3A methylation regulates expression as variation in methylation level of FAM3A was assessed in LIHC.Moreover,this over-expression of FAM3A results in poor overall survival(OS)in LIHC patients.All of these proposed that FAM3A has a role in the progression and development of LIHC.While examined association of FAM3A expression and infiltration level of CD8+T cells in LIHC patients using TIMER2 revealed that FAM3A has a positive correlation with purity in LIHC that highlights the molecular landscape.Analysis of genetic alteration revealed minute role of FAM3A in LIHC still provides valuable insight.Overall,our findings reveal that FAM3A has potential as diagnostic,therapeutic and prognostic biomarkers in LIHC.展开更多
【目的】FAM3A(family with sequence similarity 3,memberA)为新近发现的细胞因子FAM3的家族成员之一,本研究检测FAM3A的相互作用蛋白。【方法】克隆FAM3A并连接到pGB载体,转化酵母Y190宿主菌,加入含主要信号传导细胞因子人cDNA文库,...【目的】FAM3A(family with sequence similarity 3,memberA)为新近发现的细胞因子FAM3的家族成员之一,本研究检测FAM3A的相互作用蛋白。【方法】克隆FAM3A并连接到pGB载体,转化酵母Y190宿主菌,加入含主要信号传导细胞因子人cDNA文库,按标准程序进行酵母双杂交,以X-Gal显色评估活性。提取扩增阳性克隆,连接pACT2载体,测序、检索基因库确定阳性克隆基因。克隆阳性基因并与FAM3A质粒共转染酵母进一步确定相互作用。【结果】酵母双杂交检测到一个与FAM3A相互作用的阳性克隆,提取扩增后测序分析此相互作用蛋白为一睾丸特异性丝氨酸/苏氨酸激酶(TSSK4),一种与精子的发育、成熟、及生育功能有关的蛋白质.共转染pGB-FAM3A质粒与pACT2-TSSK4质粒入Y190酵母证实上述相互作用。【结论】FAM3A可能通过与TSSK4的相互作用参与精子功能与生育能力的调节。展开更多
目的:利用Ad Easy TMsystem构建携带小鼠FAM3C基因的重组腺病毒,转染至人胚肾细胞系HEK293细胞,检测外源FAM3C在细胞中的表达。方法:取小鼠肾脏提取RNA,创建c DNA文库,用PCR的方法扩增FAM3C基因,将其克隆到p EASY-1载体中,T3连接酶连接...目的:利用Ad Easy TMsystem构建携带小鼠FAM3C基因的重组腺病毒,转染至人胚肾细胞系HEK293细胞,检测外源FAM3C在细胞中的表达。方法:取小鼠肾脏提取RNA,创建c DNA文库,用PCR的方法扩增FAM3C基因,将其克隆到p EASY-1载体中,T3连接酶连接。经Bgl II单酶切后,接入p Ad Tra Ck-CMV穿梭载体,T4连接酶连接,构建重组腺病毒的穿梭质粒p Ad Tra Ck-CMV-FAM3C。将经Pme I线性化的p Ad Track-CMFAM3C电穿孔共转化入BJ5183重组细菌,获取重组腺病毒质粒Ad-FAM3C,再将经Pac I线性化的Ad-FAM3C重组病毒骨架质粒转染人胚胎肾细胞系HEK293细胞,包装并扩增病毒。用Ad-FAM3C感染小鼠肾系膜细胞,Western blot法检测FAM3C在小鼠肾系膜细胞中的表达。结果:DNA序列分析和琼脂糖凝胶电泳结果表明,已构建表达FAM3C基因的重组腺病毒,该腺病毒在体外能有效感染小鼠肾系膜细胞,且高表达FAM3C蛋白。结论:成功地构建了携带小鼠FAM3C基因的重组腺病毒,为进一步阐明FAM3C的功能及作用机制奠定实验基础。展开更多
文摘Liver hepatocellular cancer(LIHC)is positioned as the third cancer with the highest mortalities worldwide,and high mortalities are associated with late diagnosis and recurrence.This study advances bioinformatics analysis of FAM3A expression in LIHC to evaluate its potential as a prognostic,diagnostic and therapeutic biomarker.Bioinformatics tools such as UALCAN,GEPIA2,KM plotter,TIMER2 and cBioPortal are employed to conduct analysis.Initially,the expression analysis revealed up-regulation of FAM3A in LIHC based on various variables.Further,the study observed that FAM3A methylation regulates expression as variation in methylation level of FAM3A was assessed in LIHC.Moreover,this over-expression of FAM3A results in poor overall survival(OS)in LIHC patients.All of these proposed that FAM3A has a role in the progression and development of LIHC.While examined association of FAM3A expression and infiltration level of CD8+T cells in LIHC patients using TIMER2 revealed that FAM3A has a positive correlation with purity in LIHC that highlights the molecular landscape.Analysis of genetic alteration revealed minute role of FAM3A in LIHC still provides valuable insight.Overall,our findings reveal that FAM3A has potential as diagnostic,therapeutic and prognostic biomarkers in LIHC.
文摘【目的】FAM3A(family with sequence similarity 3,memberA)为新近发现的细胞因子FAM3的家族成员之一,本研究检测FAM3A的相互作用蛋白。【方法】克隆FAM3A并连接到pGB载体,转化酵母Y190宿主菌,加入含主要信号传导细胞因子人cDNA文库,按标准程序进行酵母双杂交,以X-Gal显色评估活性。提取扩增阳性克隆,连接pACT2载体,测序、检索基因库确定阳性克隆基因。克隆阳性基因并与FAM3A质粒共转染酵母进一步确定相互作用。【结果】酵母双杂交检测到一个与FAM3A相互作用的阳性克隆,提取扩增后测序分析此相互作用蛋白为一睾丸特异性丝氨酸/苏氨酸激酶(TSSK4),一种与精子的发育、成熟、及生育功能有关的蛋白质.共转染pGB-FAM3A质粒与pACT2-TSSK4质粒入Y190酵母证实上述相互作用。【结论】FAM3A可能通过与TSSK4的相互作用参与精子功能与生育能力的调节。
文摘目的:利用Ad Easy TMsystem构建携带小鼠FAM3C基因的重组腺病毒,转染至人胚肾细胞系HEK293细胞,检测外源FAM3C在细胞中的表达。方法:取小鼠肾脏提取RNA,创建c DNA文库,用PCR的方法扩增FAM3C基因,将其克隆到p EASY-1载体中,T3连接酶连接。经Bgl II单酶切后,接入p Ad Tra Ck-CMV穿梭载体,T4连接酶连接,构建重组腺病毒的穿梭质粒p Ad Tra Ck-CMV-FAM3C。将经Pme I线性化的p Ad Track-CMFAM3C电穿孔共转化入BJ5183重组细菌,获取重组腺病毒质粒Ad-FAM3C,再将经Pac I线性化的Ad-FAM3C重组病毒骨架质粒转染人胚胎肾细胞系HEK293细胞,包装并扩增病毒。用Ad-FAM3C感染小鼠肾系膜细胞,Western blot法检测FAM3C在小鼠肾系膜细胞中的表达。结果:DNA序列分析和琼脂糖凝胶电泳结果表明,已构建表达FAM3C基因的重组腺病毒,该腺病毒在体外能有效感染小鼠肾系膜细胞,且高表达FAM3C蛋白。结论:成功地构建了携带小鼠FAM3C基因的重组腺病毒,为进一步阐明FAM3C的功能及作用机制奠定实验基础。
