To construct a directional cDNA library from Chinese giant salamander Andrias davidianus liver by SMART(switching mechanism at 5′ end of RNA transcript)technique, we purified the mRNA from Andrias davidianus liver an...To construct a directional cDNA library from Chinese giant salamander Andrias davidianus liver by SMART(switching mechanism at 5′ end of RNA transcript)technique, we purified the mRNA from Andrias davidianus liver and the first strand cDNA was synthesized through reverse transcription by using a modified oligo(dT)primer(contained sfi ⅠB site). We used the SMART oligonucleotide (contained sfi ⅠA site) as a template so that the first strand cDNA could be extended over the 5′ end of mRNA. The double strand cDNA was amplified by LD PCR (long distance PCR) with the above two primers and then digested by sfi Ⅰ (ⅠA and ⅠB) restriction enzyme. After cDNA fractionation through CHROMA SPIN column, the double strand cDNA was ligated into the sfi Ⅰ digested λtripIEx2 vector and then the recombinant DNA was packaged in vitro . The content of the unamplified Andrias davidianus liver cDNA library is 1 5×10 6 in which the percentage of recombinant clones is about 98 9%. The titer of the amplified cDNA library is 1 0×10 10 pfu/ml and the average exogenous inserts of the recombinants is 1 25 kb. These results show that the Andrias davidianus liver cDNA library has excellent quality.展开更多
垂体瘤是常见的、慢性的、复杂性的颅内肿瘤,是一种多病因、多过程、多后果的全身性疾病。传统的单因素模型很难阐明垂体瘤的分子机制,而随着多组学和系统生物学的迅速发展,传统的单因素策略逐渐转变为多因素系统策略,更有利于垂体瘤的...垂体瘤是常见的、慢性的、复杂性的颅内肿瘤,是一种多病因、多过程、多后果的全身性疾病。传统的单因素模型很难阐明垂体瘤的分子机制,而随着多组学和系统生物学的迅速发展,传统的单因素策略逐渐转变为多因素系统策略,更有利于垂体瘤的预测、预防和治疗。在垂体瘤的发生发展过程中,基因组、转录组、蛋白质组、肽组、代谢组和影像组等水平的一系列分子改变均发挥了重要作用,并相互关联形成了复杂的分子网络系统。此外,多组学的重心正从结构基因组学转向表型组学(蛋白质组学和代谢组学)。在垂体瘤的多组学系统策略中,基于质谱分析的蛋白质组学和蛋白质存在形式的研究(proteoformics)起着核心作用。本文综述了多组学、基于多组学的分子通路网络、基于分子通路网络的生物标志物和治疗靶点的研究现状,并对垂体瘤的预测、预防和个体化医学(predictive,preventive and personalized medicine,PPPM)进行了展望。展开更多
文摘To construct a directional cDNA library from Chinese giant salamander Andrias davidianus liver by SMART(switching mechanism at 5′ end of RNA transcript)technique, we purified the mRNA from Andrias davidianus liver and the first strand cDNA was synthesized through reverse transcription by using a modified oligo(dT)primer(contained sfi ⅠB site). We used the SMART oligonucleotide (contained sfi ⅠA site) as a template so that the first strand cDNA could be extended over the 5′ end of mRNA. The double strand cDNA was amplified by LD PCR (long distance PCR) with the above two primers and then digested by sfi Ⅰ (ⅠA and ⅠB) restriction enzyme. After cDNA fractionation through CHROMA SPIN column, the double strand cDNA was ligated into the sfi Ⅰ digested λtripIEx2 vector and then the recombinant DNA was packaged in vitro . The content of the unamplified Andrias davidianus liver cDNA library is 1 5×10 6 in which the percentage of recombinant clones is about 98 9%. The titer of the amplified cDNA library is 1 0×10 10 pfu/ml and the average exogenous inserts of the recombinants is 1 25 kb. These results show that the Andrias davidianus liver cDNA library has excellent quality.
文摘垂体瘤是常见的、慢性的、复杂性的颅内肿瘤,是一种多病因、多过程、多后果的全身性疾病。传统的单因素模型很难阐明垂体瘤的分子机制,而随着多组学和系统生物学的迅速发展,传统的单因素策略逐渐转变为多因素系统策略,更有利于垂体瘤的预测、预防和治疗。在垂体瘤的发生发展过程中,基因组、转录组、蛋白质组、肽组、代谢组和影像组等水平的一系列分子改变均发挥了重要作用,并相互关联形成了复杂的分子网络系统。此外,多组学的重心正从结构基因组学转向表型组学(蛋白质组学和代谢组学)。在垂体瘤的多组学系统策略中,基于质谱分析的蛋白质组学和蛋白质存在形式的研究(proteoformics)起着核心作用。本文综述了多组学、基于多组学的分子通路网络、基于分子通路网络的生物标志物和治疗靶点的研究现状,并对垂体瘤的预测、预防和个体化医学(predictive,preventive and personalized medicine,PPPM)进行了展望。
