NASBA的简要过程

参考文献

NASBA

全称：nucleic acid sequence-based amplification ,依赖核酸序列的扩增技术

依赖核酸序列的扩增技术,是一种扩增RNA的新技术,是由一对引物介导的、连续均一的、体外特异核苷酸序列等温扩增的酶促反应过程.反应在42℃进行,可以在2h左右将模板RNA扩增约109～12倍,不需特殊的仪器.NASBA已经广泛应用于细菌、病毒等多种病原微生物的检测.

最早在1991年，被 J Compton 定义为：a primer-dependent technology that can be used for the continuous amplification of nucleic acids in a single mixture at one temperature

该技术一出现就被用于疾病的快速诊断和病人血清中HIV-1的定量检测。

尽管RNA的扩增也可以使用反转录PCR技术（通过反转录形成单链DNA模板),NASBA相比则有自己的优势：可以在相对恒温的条件下进行（一般恒温为41摄氏度）。

该技术用于医学诊断，相对传统的PCR技术更为稳定，准确。

NASBA的简要过程

NASBA的简要过程 如下：

1.RNA模板链进入反应混合物后，第一个引物首先与模板链的3'端结束

2. 反转录酶，合成反义，补偿的DNA链

3. RNA 酶H，分解破坏RNA模板链（仅分解与DNA杂交结合的RNA链，不分解单链RNA）

4.第二个引物与DNA的5‘端结合

5.T7 RNA polymerase 合成RNA补偿链，并加入到步骤1中，使反应可以循环进行。

NASBA技术已经被用于多个单链RNA基因组的病原性病毒的快速检测试验中，e.g. influenza A, foot-and-mouth disease virus, severe acute respiratory syndrome (SARS)-associated coronavirus, Human bocavirus(HBoV) and also parasites like Trypanosoma brucei.

参考文献

1Compton, J (1991). "Nucleic acid sequence-based amplification". Nature350 (6313): 91–2. doi:10.1038/350091a0. PMID1706072.^Kievits, T; Van Gemen, B; Van Strijp, D; Schukkink, R; Dircks, M; Adriaanse, H; Malek, L; Sooknanan, R et al. (1991).

"NASBA isothermal enzymatic in vitro nucleic acid amplification optimized for the diagnosis of HIV-1 infection". Journal of virological methods35 (3): 273–86. PMID1726172.^Schneider, P; Wolters, L; Schoone, G; Schallig, H; Sillekens, P; Hermsen, R; Sauerwein, R (2005).

"Real-time nucleic acid sequence-based amplification is more convenient than real-time PCR for quantification of Plasmodium falciparum". Journal of clinical microbiology43 (1): 402–5. doi:10.1128/JCM.43.1.402-405.2005. PMC540116. PMID15635001.Collins, RA; Ko, LS; So, KL; Ellis, T; Lau, LT; Yu, AC (2002).

"Detection of highly pathogenic and low pathogenic avian influenza subtype H5 (Eurasian lineage) using NASBA". Journal of virological methods103 (2): 213–25. PMID12008015.^Collins, RA; Ko, LS; Fung, KY; Lau, LT; Xing, J; Yu, AC (2002). "A method to detect major serotypes of foot-and-mouth disease virus". Biochemical and biophysical research communications297 (2): 267–74. PMID12237113.^Keightley, MC; Sillekens, P; Schippers, W; Rinaldo, C; George, KS (2005).

"Real-time NASBA detection of SARS-associated coronavirus and comparison with real-time reverse transcription-PCR". Journal of medical virology77 (4): 602–8. doi:10.1002/jmv.20498. PMID16254971.^B�hmer, A; Schildgen, V; L�sebrink, J; Ziegler, S; Tillmann, RL; Kleines, M; Schildgen, O (2009).

"Novel application for isothermal nucleic acid sequence-based amplification (NASBA)". Journal of virological methods158 (1-2): 199–201. doi:10.1016/j.jviromet.2009.02.010. PMID19428591.^Mugasa, CM; Laurent, T; Schoone, GJ; Kager, PA; Lubega, GW; Schallig, HD (2009).

"Nucleic acid sequence-based amplification with oligochromatography for detection of Trypanosoma brucei in clinical samples". Journal of clinical microbiology47 (3): 630–5. doi:10.1128/JCM.01430-08. PMC2650916. PMID19116352.