Objective:To investigate the effect of utilizing a molecular partner on high-level expression of Mxisca domestica(M.domestica) cecropin in Escherichia coli(E.coli) and to identify the expressed products.Methods:The ge...Objective:To investigate the effect of utilizing a molecular partner on high-level expression of Mxisca domestica(M.domestica) cecropin in Escherichia coli(E.coli) and to identify the expressed products.Methods:The genomic sequence of M.domestica cecropin A(MC) and M. domestica ubiquitin(UBI) were searched from Cenbank and amplified by reverse transcriptase polymerase chain reaction(RT-PCR).Two expression plasmids,pET32a-MC and pET32a-UBI-MC, were constructed and transferred into E.coli and were then induced by Isopropylβ-D-1- Thiogalactopyranoside(IPTG).The expression of the fusion proteins Trx-MC and Trx-UBI-MC was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Fusion protein Trx-MC was verified by Western blot analysis.The bactericidal activity of the purified MC was quantitatively determined using E.coli BL21(DE3).Results:The result showed that the fusion proteins were successively expressed in E.coli BL21 cells.A band at the expected position of 24 kDa representing the Trx-MC target protein was positivelystained,and the band at 4 kDa representing the hydrolysis of mature MC protein was also observed at the expected position. The expression levels of Trx-UBI-MC were higher than that of Trx-MC in E.coli.MC exhibited antimicrobial activity.Conclusions:With high-level expression of housefly cecropin A in E.coli using a fusion protein,MC exhibited antimicrobial activity.展开更多
c-ros致癌基因1(ROS1)重排(融合)已被广泛证实是非小细胞肺癌(NSCLC)发生的驱动因素,众多临床研究和临床实践结果证实克唑替尼显示出持久的临床获益。克唑替尼在中国已经纳入医保,是中国ROS1重排阳性NSCLC患者一线治疗的首选药物。目前...c-ros致癌基因1(ROS1)重排(融合)已被广泛证实是非小细胞肺癌(NSCLC)发生的驱动因素,众多临床研究和临床实践结果证实克唑替尼显示出持久的临床获益。克唑替尼在中国已经纳入医保,是中国ROS1重排阳性NSCLC患者一线治疗的首选药物。目前在NSCLC中至少发现了24种基因可作为ROS1的融合伴侣,不同ROS1融合突变的患者使用克唑替尼的疗效差异巨大,融合伴侣的种类可能是其重要的影响因素。目前很少有研究探讨不同融合伴侣与克唑替尼疗效的相关性。本文通过检索PubMed、ScienceDirect等文献数据库及TCGA、COSMIC和My Cancer Genome等肿瘤基因突变公共数据库,对克唑替尼治疗不同ROS1融合伴侣患者的现有研究成果和数据作一综述性分析,旨在为克唑替尼治疗不同ROS1融合伴侣NSCLC的预后判断提供依据。展开更多
基金supported by grants from National Natural Science Foundation of China(No.30972566,3087198)
文摘Objective:To investigate the effect of utilizing a molecular partner on high-level expression of Mxisca domestica(M.domestica) cecropin in Escherichia coli(E.coli) and to identify the expressed products.Methods:The genomic sequence of M.domestica cecropin A(MC) and M. domestica ubiquitin(UBI) were searched from Cenbank and amplified by reverse transcriptase polymerase chain reaction(RT-PCR).Two expression plasmids,pET32a-MC and pET32a-UBI-MC, were constructed and transferred into E.coli and were then induced by Isopropylβ-D-1- Thiogalactopyranoside(IPTG).The expression of the fusion proteins Trx-MC and Trx-UBI-MC was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Fusion protein Trx-MC was verified by Western blot analysis.The bactericidal activity of the purified MC was quantitatively determined using E.coli BL21(DE3).Results:The result showed that the fusion proteins were successively expressed in E.coli BL21 cells.A band at the expected position of 24 kDa representing the Trx-MC target protein was positivelystained,and the band at 4 kDa representing the hydrolysis of mature MC protein was also observed at the expected position. The expression levels of Trx-UBI-MC were higher than that of Trx-MC in E.coli.MC exhibited antimicrobial activity.Conclusions:With high-level expression of housefly cecropin A in E.coli using a fusion protein,MC exhibited antimicrobial activity.
文摘c-ros致癌基因1(ROS1)重排(融合)已被广泛证实是非小细胞肺癌(NSCLC)发生的驱动因素,众多临床研究和临床实践结果证实克唑替尼显示出持久的临床获益。克唑替尼在中国已经纳入医保,是中国ROS1重排阳性NSCLC患者一线治疗的首选药物。目前在NSCLC中至少发现了24种基因可作为ROS1的融合伴侣,不同ROS1融合突变的患者使用克唑替尼的疗效差异巨大,融合伴侣的种类可能是其重要的影响因素。目前很少有研究探讨不同融合伴侣与克唑替尼疗效的相关性。本文通过检索PubMed、ScienceDirect等文献数据库及TCGA、COSMIC和My Cancer Genome等肿瘤基因突变公共数据库,对克唑替尼治疗不同ROS1融合伴侣患者的现有研究成果和数据作一综述性分析,旨在为克唑替尼治疗不同ROS1融合伴侣NSCLC的预后判断提供依据。
