用于RNA模板特异性PCR的非基因组寡核苷酸标签序列的生成
Generation of non-genomic oligonucleotide tag sequences for RNA template-specific PCR.
作者信息
Pinto Fernando Lopes, Svensson Håkan, Lindblad Peter
机构信息
Department of Physiological Botany, The Angström Laboratories, Uppsala University, Box 523, SE-75120, Uppsala, Sweden.
出版信息
BMC Biotechnol. 2006 Jul 5;6:31. doi: 10.1186/1472-6750-6-31.
BACKGROUND
In order to overcome genomic DNA contamination in transcriptional studies, reverse template-specific polymerase chain reaction, a modification of reverse transcriptase polymerase chain reaction, is used. The possibility of using tags whose sequences are not found in the genome further improves reverse specific polymerase chain reaction experiments. Given the absence of software available to produce genome suitable tags, a simple tool to fulfill such need was developed.
RESULTS
The program was developed in Perl, with separate use of the basic local alignment search tool, making the tool platform independent (known to run on Windows XP and Linux). In order to test the performance of the generated tags, several molecular experiments were performed. The results show that Tagenerator is capable of generating tags with good priming properties, which will deliberately not result in PCR amplification of genomic DNA.
CONCLUSION
The program Tagenerator is capable of generating tag sequences that combine genome absence with good priming properties for RT-PCR based experiments, circumventing the effects of genomic DNA contamination in an RNA sample.
背景
为了克服转录研究中的基因组DNA污染问题，人们采用了反向模板特异性聚合酶链反应，这是逆转录酶聚合酶链反应的一种改良方法。使用基因组中不存在其序列的标签进一步改进了反向特异性聚合酶链反应实验。鉴于缺乏可用于生成适合基因组的标签的软件，开发了一种满足此类需求的简单工具。
结果
该程序用Perl语言开发，单独使用基本局部比对搜索工具，使该工具具有平台独立性（已知可在Windows XP和Linux上运行）。为了测试生成的标签的性能，进行了几项分子实验。结果表明，Tagenerator能够生成具有良好引物特性的标签，这些标签不会导致基因组DNA的PCR扩增。
结论
Tagenerator程序能够生成结合了基因组不存在特性和良好引物特性的标签序列，用于基于RT-PCR的实验，规避RNA样本中基因组DNA污染的影响。
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