Bingkun Tian, Hongjing Wu, Rongrong Wang, Hong Chen, Haixing Li
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引用次数: 0
Abstract
As a tool for acquiring uncharacterized genomic DNA adjacent to characterized DNA, genome-walking is integral to bioscience-related research works. Herein, a new genome-walking tool known as N7-ended walker PCR (polymerase chain reaction) is presented. The key aspect for this method is the use of a degenerate walker primer in secondary/tertiary PCR. The 7 nt 5' tail of this primer completely degenerates to N7 relative to its corresponding primary walker primer. The degeneracy reduces the efficiency of annealing this primer to its predecessor site. Clearly, primary nontarget DNA defined by the primary walker primer prefers to form a hairpin structure via the inverted ends rather than hybridizing with the degenerate primer. As a result, N7-ended walker PCR achieves genome-walking by selectively boosting the DNA of interest. The feasibility of the N7-ended walker PCR method was proven by acquiring uncharacterized DNAs flanking several characterized DNAs.
作为获取与特征 DNA 相邻的未表征基因组 DNA 的工具，基因组漫步是生物科学相关研究工作不可或缺的一部分。本文介绍了一种新的基因组漫步工具，即 N7 端漫步 PCR（聚合酶链反应）。这种方法的关键在于在二级/三级 PCR 中使用退化的步行者引物。该引物的 7 nt 5' 尾部相对于其相应的主引物完全退化为 N7。这种退化降低了该引物退火到其前导位点的效率。显然，由主走行引物定义的原生非目标 DNA 更喜欢通过倒置末端形成发夹结构，而不是与退化引物杂交。因此，N7 端走行 PCR 通过选择性地增强感兴趣的 DNA 来实现基因组走行。N7-ended walker PCR 方法的可行性通过获取几个特征 DNA 侧面的未特征 DNA 得到了证明。
期刊介绍：
Biochemical Genetics welcomes original manuscripts that address and test clear scientific hypotheses, are directed to a broad scientific audience, and clearly contribute to the advancement of the field through the use of sound sampling or experimental design, reliable analytical methodologies and robust statistical analyses.
Although studies focusing on particular regions and target organisms are welcome, it is not the journal’s goal to publish essentially descriptive studies that provide results with narrow applicability, or are based on very small samples or pseudoreplication.
Rather, Biochemical Genetics welcomes review articles that go beyond summarizing previous publications and create added value through the systematic analysis and critique of the current state of knowledge or by conducting meta-analyses.
Methodological articles are also within the scope of Biological Genetics, particularly when new laboratory techniques or computational approaches are fully described and thoroughly compared with the existing benchmark methods.
Biochemical Genetics welcomes articles on the following topics: Genomics; Proteomics; Population genetics; Phylogenetics; Metagenomics; Microbial genetics; Genetics and evolution of wild and cultivated plants; Animal genetics and evolution; Human genetics and evolution; Genetic disorders; Genetic markers of diseases; Gene technology and therapy; Experimental and analytical methods; Statistical and computational methods.
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