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| A Visualized Closed-tube Detection Method for Loop-mediated Isothermal Amplification Based on Nucleic Acid Molecular Light Switch |
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| DOI： |
| KeyWord:LAMP isothermal amplification ?［Ru（bpy）2（dppz）］2+ ?visual detection ?S. aureus |
| Author | Institution |
| WANG Fang，DONG Jing，LI Yan-ni，XU Qin-feng |
School of Food and Bioengineering，Shaanxi University of Science and Technology，Xi′an ，China |
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| Abstract: |
| Compared with polymerase chain reaction（PCR） nucleic acid amplification，loop-mediated isothermal amplification（LAMP） does not require thermal cycling instruments，which is more suitable for rapid on-site detection．However，the small Stoke′s shift of currently used organic fluorescent dye for visualized LAMP leads to insufficient color differentiation．In this study，the nucleic acid molecule“light switch”［Ru（bpy）2（dppz）］2+ was exploited as a LAMP fluorescent dye to establish a rapid visualized closed-tube method for LAMP detection．Visualization detection requires a high concentration of dye，but its adding before amplification reaction will strongly inhibit LAMP reaction，while its adding after reaction by opening the lid will lead to a serious aerosol cross contamination．Therefore，wax was chosen to seal the［Ru（bpy）2（dppz）］2+ at the bottom of the tube，and the LAMP amplification products were visually detected by using the strong red fluorescence emission of ［Ru（bpy）2（dppz）］2+ in the presence of double-strand DNA．The S.aureus DNA could be detected as low as 20 copies/reaction with strong specificity，and the whole visual detection process could be completed within 1 h．It could be widely used for the rapid on-site nucleic acid detection in the fields of environmental detection，food safety and clinical diagnosis． |
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