| 高GC含量DNA序列PCR扩增的引物设计 |
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| 引用本文： | 高梅,;刘树人,;李强,;张凌峰. 高GC含量DNA序列PCR扩增的引物设计[J]. 华南国防医学杂志, 2014, 0(7): 633-638 |
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| 作者姓名： | 高梅, 刘树人, 李强, 张凌峰 |
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| 作者单位： | [1]解放军458医院检验科,广东广州510602; [2]解放军458医院传染科,广东广州510602; [3]解放军458医院血库,广东广州510602; [4]南方医科大学南方医院检验科,广东广州510602; |
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| 基金项目： | 全军医学科学技术研究“十一五”计划保健专项课题（08BJZ07） |
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| 摘 要： | 目的建立一种高GC含量DNA序列PCR扩增的引物设计方法。方法共设计了15对高GC含量（66.0%~84.0%）DNA序列PCR扩增引物,引物Tm值均≥79.7℃,引物Tm差值（△Tm）均≤1℃。对本研究所设计引物及相关参考文献中部分引物的参数进行统计学分析。另外,基于本研究方法,设计了26对引物,以验证该方法在非高GC含量（35.2%~53.5%）DNA序列PCR扩增中的实用性。结果采用本研究方法,所有15对高GC含量DNA序列均成功扩增;统计结果显示,Tm值和△Tm是影响高GC含量DNA序列扩增的主要因素。结论设计具有较高Tm值和较低△Tm的引物可有效的解决高GC含量DNA序列的PCR扩增问题。另外,在较高的退火温度下,引物或DNA序列的二级结构无法形成。
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| 关 键 词： | 聚合酶链反应 高GC含量 引物设计 |
Primer Design Method for PCR of GC-rich DNA Tamplates |
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| Affiliation: | GAO Mei , LIU Shu-ren , LI Qiang , ZHANG Ling-feng( Clinical Laboratory, No. 458 Hospital of the People＇s Liberation Army, Guangzhou Guangdong 510602, China) |
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| Abstract: | Objective To establish a primer design method for amplification of GC-rich DNA sequences.Methods A group of 15 pairs of primers with higher Tm （≥79.7℃） and lower△Tm （≤11 ℃） were designed to amplify GC-rich sequences （66.0%-84.0%）. The statistical analysis of primer parameters and GC content of PCR products was per formed and compared with literatures. A group of 26 pairs of primers were designed to test the applicability of this primer designing method in amplifications of non-GCrich sequences （35.2%-53.5%）.Results All the DNA sequences in this study were successfully amplified. Tm and ATm were the main factors influencing amplifications.Conclusions This prim- er designing method offered a perfect tool for amplification of GC rich sequences. It proves that the secondary structures cannot be formed at higher annealing temperature conditions. |
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| Keywords: | Polymerase chain reaction GC-rich sequences Primer design |
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