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et重组
相关语句
  et recombination
     Red/ET Recombination and its Biomedical Applications
     Red/ET重组及其在生物医学中的应用
短句来源
     ET Recombination: new options for cloning DNA in Escherichia coli
     ET重组:Escherichia coli中最新分子克隆方法
短句来源
     The Application of Red/ET Recombination to High Efficient Gene-targeting Vector Construction
     Red/ET重组在基因打靶载体快速构建中的应用
短句来源
     Red/ET recombination, a powerful homologous recombination system based on the Red operon of λ phage or RecE/RecT from Rac phage, provides an innovative approach for DNA engineering.
     通过应用Rac噬菌体的RecE RecT和λ噬菌体的RedαRedβ系统而建立的DNA工程平台———Red ET重组,是一种不依赖于限制性内切酶的分子克隆新技术。
短句来源
     Red/ET recombination has the advantage of getting longer homology regions without mutation,which makes it a new and reliable alternative to the construction of a targeting vector today.
     因此Red/ET重组为构建打靶载体提供了一种新的可靠的方法。
短句来源
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  相似匹配句对
     et.
     et.
短句来源
     Wu et Y.
     W u et Y.
短句来源
     The recombinant plasmid expressed well in E.
     从重组E.
短句来源
     Red/ET Recombination and its Biomedical Applications
     Red/ET重组及其在生物医学中的应用
短句来源
     Rebuild CRM
     重组CRM
短句来源
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  et recombination
Because the BAC that we used contained several genes, we trimmed it by ET recombination12 to carry Hc only.
      
By using Red/ET recombination technology, we were able to circumvent many of the present limitations in the engineering of polyketide systems.
      
To accomplish this goal, an approach using Red/ET recombination with gene complementation was developed.
      
The resulting strain was used as the host for Red/ET recombination.
      


In Escherichia coli most recombinations depend on intracellular RecA protein. RecBCD degrades double-stranded DNA, so only circular dsDNA with long homology as targeting cassettes can carry out recombination in vivo. Recently, highly efficient phage-based E.coli homologous recombination systems including Rac-encoded RecET system and λ-encoded Red system have been developed. These phage systems can catalyze efficient recombination with very short homology segments. Mutation such as insertion, deletion, substitution,...

In Escherichia coli most recombinations depend on intracellular RecA protein. RecBCD degrades double-stranded DNA, so only circular dsDNA with long homology as targeting cassettes can carry out recombination in vivo. Recently, highly efficient phage-based E.coli homologous recombination systems including Rac-encoded RecET system and λ-encoded Red system have been developed. These phage systems can catalyze efficient recombination with very short homology segments. Mutation such as insertion, deletion, substitution, single-base changes and in vivo cloning by gap repair can be introduced into E.coli chromosome DNA or eukaryotic genomics DNA cloned in BACs or PACs. Importantly they need no restriction enzymes and DNA ligase, eliminate many of the time-consuming in vitro steps of construction. Recombineering offers a powerful new tool for functional genomics.

大肠杆菌的同源重组依靠内源性的RecA蛋白。RecBCD降解双链线性DNA分子,因此必须构建环状质粒打靶载体才能完成体内重组,操作过程繁琐,所需同源臂长。最近建立起的依赖于Rac噬菌体的ET重组系统和基于λ噬菌体的Red重组系统,可有效利用线性DNA片段作为打靶分子,对大肠杆菌染色体DNA和BAC、PAC载体中包含的真核细胞基因组DNA进行基因敲除、敲入、替换、单碱基突变及体内基因克隆等修饰。这2种系统重组效率高,用PCR方法便可合成双链线性DNA打靶分子,不需要限制酶和连接酶,操作过程简单、精确、快速、经济,大大缩短了构建打靶载体的时间,成为功能基因组研究的有力工具。

Red/ET recombination, a powerful homologous recombination system based on the Red operon of λ phage or RecE/RecT from Rac phage, provides an innovative approach for DNA engineering. Deletion, insertion and mutation can be quickly and precisely performed on the target gene mediated by Red/ET recombination with PCR derived DNA fragments or oligonucleotides. This technical platform has extensive applications in biomedical field including bacterial artificial chromosome modification, gene knock-out construction...

Red/ET recombination, a powerful homologous recombination system based on the Red operon of λ phage or RecE/RecT from Rac phage, provides an innovative approach for DNA engineering. Deletion, insertion and mutation can be quickly and precisely performed on the target gene mediated by Red/ET recombination with PCR derived DNA fragments or oligonucleotides. This technical platform has extensive applications in biomedical field including bacterial artificial chromosome modification, gene knock-out construction and genetic modification of E.coli strains as well as some other kinds of microorganisms. Recently, Red/ET recombination was improved in several aspects so that it becomes more powerful and maneuverable. The characteristic and development of Red/ET recombination and its biomedical applications were described in this review.

通过应用Rac噬菌体的RecE RecT和λ噬菌体的RedαRedβ系统而建立的DNA工程平台———Red ET重组,是一种不依赖于限制性内切酶的分子克隆新技术。运用该技术能够介导PCR产物或寡核苷酸对目标基因进行剪切、插入、融合及突变等多种操作,在生物医学领域里具有广阔的应用前景,尤其在基因组功能研究中对BACs、PACs和细菌染色体的打靶修饰以及基因敲除动物DNA靶分子的快速构建等方面最有效。随着Red ET重组的推广与应用,该技术本身也在不断被改进,在工作效率得到显著提高的同时,其操作也变得更加简单、省时、省力。结合自身的一些研究结果,对Red ET重组的技术特点、发展现状和在生物医学中的应用进行了详细阐述。

A rapid and high efficient working system for gene-targeting vector construction was developed by using Red/ET recombination.Mediated by Red/ET recombination,the objective genomic DNA was first subcloned into the targeting vector.After insertion of a PCR amplified selectable marker gene flanked with short homology arms into the targeted position,a conventional gene knock-out targeting vector was then constructed.For conditional gene knock-out targeting vector construction,with the co-operation of Cre-loxP site-specific...

A rapid and high efficient working system for gene-targeting vector construction was developed by using Red/ET recombination.Mediated by Red/ET recombination,the objective genomic DNA was first subcloned into the targeting vector.After insertion of a PCR amplified selectable marker gene flanked with short homology arms into the targeted position,a conventional gene knock-out targeting vector was then constructed.For conditional gene knock-out targeting vector construction,with the co-operation of Cre-loxP site-specific recombination,two rounds of Red/ET recombination was just needed.Being different from PCR and endonuclease-based gene-targeting vector construction,the homologous regions used for gene targeting can be chosen as long as possible.Furthermore,no enzyme digestion,ligation and sequencing identification were involved,so that it is very efficient and labor-saving.Several different gene-targeting vectors were successfully constructed by using this system.The establishment of this working system will accelerate the gene function studies in the post-genome stage.

通过合理应用Red/ET重组技术实现基因打靶载体的快速构建。在Red/ET重组介导下,首先从基因组DNA中将靶基因片段亚克隆至打靶质粒载体中,随后将两端带有50 bp同源臂的抗性筛选基因插入并替换靶基因上的目标序列,如此两步操作即可完成一个传统型基因敲除打靶载体的构建;结合Cre-loxP系统,在传统型基因敲除打靶载体的基础上,经过再一轮的Red/ET重组就能够成功实现条件性基因敲除打靶载体的构建。整个实验过程不需要PCR扩增长、短臂序列,也不涉及酶切、连接反应,因此,不仅省时、省力,而且所构建的基因打靶载体序列准确,无突变。实验方法的建立为加速后基因组时代的基因功能研究提供了一条捷径。

 
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