Jang MinJu, Kim Sanghyo
Department of Bionanotechnology, Gachon University, Seongnam, 13120 Republic of Korea.
Biochip J. 2022;16(3):326-333. doi: 10.1007/s13206-022-00070-3. Epub 2022 Jul 27.
Loop-mediated isothermal amplification (LAMP) may be used in molecular and point-of-care diagnostics for pathogen detection. The amplification occurs under isothermal conditions using up to six primers. However, non-specific amplification is frequently observed in LAMP. Non-specific amplification has the potential to be triggered by forward and reverse internal primers. And the relatively low reaction temperature (55-65 °C) induces the secondary structure via primer-primer interactions. Primer redesign and probe design have been recommended to solve this problem. LAMP primers have strict conditions, such as Tm, GC contents, primer dimer, and distance between primers compared to conventional PCR primers. Probe design requires specialized knowledge to have high specificity for a target. In polymerase chain reaction (PCR), some chemicals or proteins are used for improving specificity and efficiency. Therefore, we hypothesized that additives can suppress the non-specific amplification. In this study, tetramethylammonium chloride (TMAC), formamide, dimethyl sulfoxide, Tween 20, and bovine serum albumin have been used as LAMP additives. In our study, TMAC was presented as a promising additive for suppressing non-specific amplification in LAMP.
The online version contains supplementary material available at 10.1007/s13206-022-00070-3.
环介导等温扩增（LAMP）可用于分子诊断和即时诊断中的病原体检测。该扩增在等温条件下使用多达六种引物进行。然而，LAMP中经常观察到非特异性扩增。非特异性扩增有可能由正向和反向内部引物触发。并且相对较低的反应温度（55 - 65°C）通过引物 - 引物相互作用诱导二级结构。建议重新设计引物和设计探针来解决这个问题。与传统PCR引物相比，LAMP引物有严格的条件，如熔解温度、GC含量、引物二聚体以及引物之间的距离。探针设计需要专业知识以对靶标具有高特异性。在聚合酶链反应（PCR）中，一些化学物质或蛋白质用于提高特异性和效率。因此，我们假设添加剂可以抑制非特异性扩增。在本研究中，氯化铵（TMAC）、甲酰胺、二甲基亚砜、吐温20和牛血清白蛋白已被用作LAMP添加剂。在我们的研究中，TMAC被证明是一种有前景的用于抑制LAMP中非特异性扩增的添加剂。
在线版本包含可在10.1007/s13206 - 022 - 00070 - 3获取的补充材料。
