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首页> 外文期刊>Biochemistry >Polyamide nucleic acid DNA chimera lacking the phosphate backbone are novel primers for polymerase reaction catalyzed by DNA polymerases
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Polyamide nucleic acid DNA chimera lacking the phosphate backbone are novel primers for polymerase reaction catalyzed by DNA polymerases

机译：缺乏磷酸骨架的聚酰胺核酸DNA嵌合体是DNA聚合酶催化的新型聚合酶反应引物

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摘要

A peptide nucleic acid (PNA) oligomer, an analogue of DNA, was examined for its ability to function as a primer or a template to support DNA synthesis catalyzed by DNA polymerases. The analogue, (PNA)(19)-TpG-OH, comprised of 19 bases in the form of PNA followed by a dinucleotide (TpG-OH) with a single phosphate and a free 3'OH terminus, was recognized as a bona fide primer by 2 reverse transcriptases and also by the Klenow fragment of E. coli DNA polymerase I, The 21-mer PNA chimera is extended on both RNA and DNA templates by all three polymerases. The specificity of binding of the PNA chimeric primer/DNA template at the template-primer binding site of the enzyme was shown by its photo-cross-linking ability to the enzyme which could be effectively competed out by another TP but not by template or primer alone, Furthermore, the chimeric TP-enzyme covalent complex was found to be catalytically active as judged by its ability to incorporate one nucleotide onto the 3'OH terminus of the immobilized primer, PNA sequences were also recognized as template when annealed with a DNA primer. These observations are in variance with the conventional suggestion that the phosphate backbone in the duplex region in essential for recognition and binding hv DNA polymerases. The efficient extension of (PNA)(19)-TpG-OH suggests that the diameter of the duplex region of template primer rather than the phosphate backbone may be sufficient for recognition by DNA polymerases. [References: 24]
机译：检查了肽核酸（PNA）低聚物（一种DNA的类似物）的功能，以作为引物或模板来支持DNA聚合酶催化的DNA合成。公认的类似物（PNA）（19）-TpG-OH由19个PNA形式的碱基组成，其后是带有一个磷酸酯和一个游离3'OH末端的二核苷酸（TpG-OH），被认为是真正的通过2种逆转录酶以及大肠杆菌DNA聚合酶I的Klenow片段作为引物，21种PNA嵌合体在这三种聚合酶上均在RNA和DNA模板上延伸。 PNA嵌合引物/ DNA模板在酶的模板引物结合位点的结合特异性由其与酶的光交联能力显示，该酶可以通过另一种TP有效竞争，但不能通过模板或引物竞争此外，发现嵌合TP-酶共价复合物具有催化活性，这是根据其将一个核苷酸掺入固定引物3'OH末端的能力来判断的，当与DNA引物退火时，PNA序列也被认为是模板。 。这些观察结果与传统的建议不一致，即在双链体区域中的磷酸骨架对于识别和结合hv DNA聚合酶必不可少。 （PNA）（19）-TpG-OH的有效延伸表明模板引物的双链体区域而不是磷酸盐骨架的直径可能足以被DNA聚合酶识别。 [参考：24]

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