重组酶聚合酶扩增技术研究进展

doi:10.13560/j.cnki.biotech.bull.1985.2016.06.008

摘要/Abstract

摘要： 重组酶聚合酶扩增(recombinase polymerase amplification，RPA)技术是一种新兴的核酸恒温扩增技术，具有灵敏度高、特异性强、反应快速等特点。利用RPA技术可以对核酸拷贝数进行绝对定量并且可以同时检测多个靶标核酸序列，结合侧流层析试纸条或荧光信号监测装置等简易实验设备即可直接观察检测结果。就RPA技术的原理、发展、技术特点及其近年来在体外诊断、病原检测等领域的研究进展作一综述，旨在为该技术的深入研究和应用提供参考。

关键词: 重组酶聚合酶扩增技术, 核酸恒温扩增, 核酸检测

Abstract: Recombinase Polymerase Amplification(RPA)is a recently developed isothermal amplification method that offers highly sensitive and specific DNA and RNA detection. Using RPA，the initial copy number of target nucleic acid sequence from different samples can be absolutely quantified，and multiple target nucleic acid sequences can also be detected simultaneously. Combining with less complicated device such as lateral flow strips or a sequence-specific fluorescent probe，the results can be observed directly. This article summarized the fundamental principles，continuous development and technical features of RPA. In addition，research and application progress of RPA in the field of in vitro diagnostic，pathogen detection and so forth were reviewed，aiming at providing guidance for further development and application of this method.

Key words: recombinase polymerase amplification, nucleic acid isothermal amplification technologies, nucleic acid detection

景志刚, 董浩, 狄栋栋, 田莉莉, 范伟兴. 重组酶聚合酶扩增技术研究进展[J]. 生物技术通报, 2016, 32(6): 47-53.

JING Zhi-gang, DONG Hao, DI Dong-dong, TIAN Li-li, FAN Wei-xing. Research Progress on Recombinase Polymerase Amplification[J]. Biotechnology Bulletin, 2016, 32(6): 47-53.

参考文献

[1]Piepenburg O, Williams CH, Stemple DL, et al. DNA detection using recombination proteins[J]. PLoS Biology, 2006, 4(7)：e204.
[2]Kersting S, Rausch V, Bier FF, et al. Multiplex isothermal solid-phase recombinase polymerase amplification for the specific and fast DNA-based detection of three bacterial pathogens[J]. Microchimica Acta, 2014, 181(13-14)：1715-1723.
[3] Xu C, Li L, Jin W, et al. Recombinase polymerase amplification(RPA)of CaMV-35S promoter and nos terminator for rapid detection of genetically modified crops[J]. International Journal of Molecular Sciences, 2014, 15(10)：18197-18205.
[4]Shen F, Davydova EK, Du W, et al. Digital isothermal quantification of nucleic acids via simultaneous chemical initiation of recombinase polymerase amplification reactions on SlipChip[J]. Analytical Chemistry, 2011, 83(9)：3533-3540.
[5]Boyle DS, McNerney R, Teng Low H, et al. Rapid detection of Mycobacterium tuberculosis by recombinase polymerase amplification[J]. PLoS One, 2014, 9(8)：e103091.
[6]Mekuria TA, Zhang S, Eastwell KC. Rapid and sensitive detection of Little cherry virus 2 using isothermal reverse transcription-recombinase polymerase amplification[J]. Journal of Virological Methods, 2014, 205(1)：24-30.
[7]Abd El Wahed A, El-Deeb A, El-Tholoth M, et al. A portable reverse transcription recombinase polymerase amplification assay for rapid detection of foot-and-mouth disease virus[J]. PLoS One, 2013, 8(8)：e71642.
[8]Rohrman BA, Richards-Kortum RR. A paper and plastic device for performing recombinase polymerase amplification of HIV DNA[J]. Lab Chip, 2012, 12(17)：3082-3088.
[9]Wee EJ, Lau HY, Botella JR, et al. Re-purposing bridging flocculation for on-site, rapid, qualitative DNA detection in resource-poor settings[J]. Chemical Communications, 2015, 51(27)：5828-5831.
[10]Santiago-Felipe S, Tortajada-Genaro LA, Morais S, et al. One-pot isothermal DNA amplification-Hybridisation and detection by a disc-based method[J]. Sensors and Actuators B：Chemical, 2014, 204(1)：273-281.
[11]Aebischer A, Wernike K, Hoffmann B, et al. Rapid genome detection of schmallenberg virus and bovine viral diarrhea virus by use of isothermal amplification methods and high-speed real-time reverse transcriptase PCR[J]. Journal of Clinical Microbiology, 2014, 52(6)：1883-1892.
[12]Euler M, Wang Y, Heidenreich D, et al. Development of a panel of recombinase polymerase amplification assays for detection of biothreat agents[J]. Journal of Clinical Microbiology, 2013, 51(4)：1110-1117.
[13]Ahmed A, van der Linden H, Hartskeerl RA. Development of a recombinase polymerase amplification assay for the detection of pathogenic Leptospira[J]. Int J Environ Res Public Health, 2014, 11(5)：4953-4964.
[14]Xia X, Yu Y, Weidmann M, et al. Rapid detection of shrimp white spot syndrome virus by real time, isothermal recombinase polymerase amplification assay[J]. PLoS One, 2014, 9(8)：e104667.
[15]Boyle DS, Lehman DA, Lillis L, et al. Rapid detection of HIV-1 proviral DNA for early infant diagnosis using recombinase polymerase amplification[J]. MBio, 2013, 4(2)：e00135-00113.
[16]Kersting S, Rausch V, Bier FF, et al. Rapid detection of Plasmodium falciparum with isothermal recombinase polymerase amplification and lateral flow analysis[J]. Malaria Journal, 2014, 13(1)：99.
[17]Escadafal C, Faye O, Sall AA, et al. Rapid molecular assays for the detection of yellow fever virus in low-resource settings[J]. PLoS Neglected Tropical Diseases, 2014, 8(3)：e2730.
[18]Yan L, Zhou J, Zheng Y, et al. Isothermal amplified detection of DNA and RNA[J]. Molecular Biosystems, 2014, 10(5)：970-1003.
[19]Fakruddin M, Mannan KS, Chowdhury A, et al. Nucleic acid amplification：Alternative methods of polymerase chain reaction[J]. Journal of Pharmaceutical Sciences, 2013, 5(4)：245-252.
[20]Craw P, Balachandran W. Isothermal nucleic acid amplification technologies for point-of-care diagnostics：a critical review[J]. Lab Chip, 2012, 12(14)：2469-2486.
[21] Gill P, Ghaemi A. Nucleic acid isothermal amplification technol-ogies：a review[J]. Nucleosides Nucleotides Nucleic Acids, 2008, 27(3)：224-243.
[22]Zaghloul H, El-Shahat M. Recombinase polymerase amplification as a promising tool in hepatitis C virus diagnosis[J]. World Journal of Hepatology, 2014, 6(12)：916-922.
[23]Shin Y, Perera AP, Kim KW, et al. Real-time, label-free isothermal solid-phase amplification/detection(ISAD)device for rapid detection of genetic alteration in cancers[J]. Lab Chip, 2013, 13(11)：2106-2114.
[24]Loo JF, Lau PM, Ho HP, et al. An aptamer-based bio-barcode assay with isothermal recombinase polymerase amplification for cytochrome-c detection and anti-cancer drug screening[J]. Talanta, 2013, 115(4)：159-165.
[25]Santiago-Felipe S, Tortajada-Genaro LA, Puchades R, et al. Recombinase polymerase and enzyme-linked immunosorbent assay as a DNA amplification-detection strategy for food analysis[J]. Analytica Chimica Acta, 2014, 811(5)：81-87.
[26]徐潮, 李亮, 金芜军, 宛煜嵩. 荧光RPA技术检测转基因水稻科丰6号[J]. 分子植物育种, 2014, 12(5)：875-880.
[27]Liljander A, Yu M, O’Brien E, et al. Field-applicable recombinase polymerase amplification assay for rapid detection of Mycoplasma capricolum subsp. capripneumoniae[J]. Journal of Clinical Microbiology, 2015, 53(9)：2810-2815.
[28]Lillis L, Lehman D, Singhal MC, et al. Non-instrumented incubation of a recombinase polymerase amplification assay for the rapid and sensitive detection of proviral HIV-1 DNA[J]. PLoS One, 2014, 9(9)：e108189.
[29]Abd El Wahed A, Weidmann M, Hufert FT. Diagnostics-in-a-Suitcase：Development of a portable and rapid assay for the detection of the emerging avian influenza A(H7N9)virus[J]. Journal of Clinical Virology, 2015, 69(3)：16-21.
[30] Euler M, Wang Y, Otto P, et al. Recombinase polymerase amplific-ation assay for rapid detection of Francisella tularensis[J]. Journal of Clinical Microbiology, 2012, 50(7)：2234-2238.
[31]Daher RK, Stewart G, Boissinot M, et al. Isothermal recombinase polymerase amplification assay applied to the detection of group B streptococci in vaginal/anal samples[J]. Clinical Chemistry, 2014, 60(4)：660-666.
[32]Kim TH, Park J, Kim CJ, et al. Fully integrated lab-on-a-disc for nucleic acid analysis of food-borne pathogens[J]. Analytical Chemistry, 2014, 86(8)：3841-3848.
[33]Tsaloglou MN, Watson RJ, Rushworth CM, et al. Real-time microfluidic recombinase polymerase amplification for the toxin B gene of Clostridium difficile on a SlipChip platform[J]. Analyst, 2015, 140(1)：258-264.
[34]Murinda SE, Ibekwe AM, Zulkaffly S, et al. Real-time isothermal detection of Shiga toxin-producing Escherichia coli using recombinase polymerase amplification[J]. Foodborne Pathogens and Disease, 2014, 11(7)：529-536.
[35] Ahmed SA, van de Sande WW, Desnos-Ollivier M, et al. Applica-tion of isothermal amplification techniques for the identification of Madurella mycetomatis, the prevalent agent of human mycetoma[J]. Journal of Clinical Microbiology, 2015, 53(10)：3280-3285.
[36]Yehia N, Arafa AS, Abd El Wahed A, et al. Development of reverse transcription recombinase polymerase amplification assay for avian influenza H5N1 HA gene detection[J]. Journal of Virological Methods, 2015, 223：45-49.
[37]Teoh BT, Sam SS, Tan KK, et al. Early detection of dengue virus by use of reverse transcription-recombinase polymerase amplification[J]. Journal of Clinical Microbiology, 2015, 53(3)：830-837.
[38]Zhang S, Ravelonandro M, Russell P, et al. Rapid diagnostic detection of plum pox virus in Prunus plants by isothermal Amplify-RP? using reverse transcription-recombinase polymerase amplification[J]. Journal of Virological Methods, 2014, 207(2)：114-120.
[39]Abd El Wahed A, Patel P, Heidenreich D, et al. Reverse transcription recombinase polymerase amplification assay for the detection of middle east respiratory syndrome coronavirus[J]. PLoS Currents, 2013, 5：ecurrents. outbreaks. 62df1c7c75ffc96cd59034531e2e8364.
[40]Amer HM, Abd El Wahed A, Shalaby MA, et al. A new approach for diagnosis of bovine coronavirus using a reverse transcription recombinase polymerase amplification assay[J]. Journal of Virological Methods, 2013, 193(2)：337-340.
[41]Silva G, Bomer M, Nkere C, et al. Rapid and specific detection of Yam mosaic virus by reverse-transcription recombinase polymerase amplification[J]. Journal of Virological Methods, 2015, 15(1)：138-144.
[42]Xia X, Yu Y, Hu L, et al. Rapid detection of infectious hypodermal and hematopoietic necrosis virus(IHHNV)by real-time, isothermal recombinase polymerase amplification assay[J]. Archives of Virology, 2015, 160(4)：987-994.
[43]Krolov K, Frolova J, Tudoran O, et al. Sensitive and rapid detection of Chlamydia trachomatis by recombinase polymerase amplification directly from urine samples[J]. J Mol Diagn, 2014, 16(1)：127-135.
[44]Chao CC, Belinskaya T, Zhang Z, et al. Development of recombinase polymerase amplification assays for detection of Orientia tsutsugamushi or Rickettsia typhi[J]. PLoS Negl Trop Dis, 2015, 9(7)：e0003884.
[45]Nair G, Rebolledo M, White AC Jr, et al. Detection of entamoeba histolytica by recombinase polymerase amplification[J]. American Journal of Tropical Medicine and Hygiene, 2015, 93(3)：591-595.
[46]Crannell ZA, Cabada MM, Castellanos-Gonzalez A, et al. Recombinase polymerase amplification-based assay to diagnose giardia in stool samples[J]. American Journal of Tropical Medicine and Hygiene, 2015, 92(3)：583-587.
[47]Castellanos-Gonzalez A, Saldarriaga OA, Tartaglino L, et al. A novel molecular test to diagnose canine visceral Leishmaniasis at the point of care[J]. American Journal of Tropical Medicine and Hygiene, 2015, 3：15-0145.
[48]Rosser A, Rollinson D, Forrest M, et al. Isothermal recombinase polymerase amplification(RPA)of Schistosoma haematobium DNA and oligochromatographic lateral flow detection[J]. Parasites & Vectors, 2015, 8：446.