Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

LAMP Assay Optimization

MK Mehran Khan
RW Rongbo Wang
BL Benjin Li
PL Peiqing Liu
QW Qiyong Weng
QC Qinghe Chen
request Request a Protocol
ask Ask a question
Favorite

The LAMP assay was optimized using different LAMP primer concentrations [F3/B3 (0.1–0.5 μM) and FIP/BIP (0.8–2.4 μM)], durations (30–70 min), temperatures (57–67°C), and reagents [dNTPs (0.8–2.0 mM), MgSO4 (0.5–3.0 mM), and betaine (1.5–4 M)]. The LAMP system used in this study consisted of 12.5 μL LAMP buffer, 4 μL primer mix (F3/B3 and FIP/BIP), 1 μL calcein-MnCl2, 1 μL template DNA, 1 μL Bst DNA polymerase (8000U, New England Biolabs), and sterile double-distilled water for a final volume of 25 μL. The LAMP reaction was conducted in a thermal water bath by adjustment to the optimized temperature of 63°C for 60 min. The amplified LAMP products were further observed on 2% stained agarose gel with ethidium bromide to confirm amplification. All reactions were repeated at least three times.

Copyright and License information:  ©2018 Khan, Wang, Li, Liu, Weng and Chen.
This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

post Post a Question
0 Q&A