Journal: BMC Infectious Diseases
Article Title: Development and evaluation of reverse transcription-loop-mediated isothermal amplification assay for rapid detection of enterovirus 71
Figure Lengend Snippet: The optimal condition of RT-LAMP for the detection of EV71 . (2A) The optimal temperature of RT-LAMP assay is monitored by agarose gel analysis. M: Marker, 100-bp DNA ladder (Sigma, USA); Lanes 1-6 are reaction temperatures at 60, 61, 62, 63, 64 and 65°C, respectively. (2B) The reaction time of RT-LAMP. Lanes 1-5 are reaction time of 30, 40, 50, 60 and 70 min, respectively. (2C) The concentration of magnesium. Lanes 1-5 are magnesium at the concentrations of 0, 1, 2, 2.8 and 3.6 mM, respectively. (2D) The concentration of betaine. Lanes 1-4 are the betaine concentrations of 0, 0.5, 1.0 and 1.5 M, respectively.
Article Snippet: Reaction system of RT-LAMP The RT-LAMP reaction was performed in a 25 μl of reaction mixture containing 0.8 μM (each) FIP and BIP inner primers, 0.2 μM forward outer primer (F3) and backward outer primer (B3), 0.4 μM LF and LB (each), 0.4 mM deoxynucleoside triphosphates (dNTPs), 1 M betaine (Sigma, USA), 20 mM Tris-HCl (pH 8.8), 10 mM KCl, 10 mM (NH4 )2 SO4 , 2.8 mM MgSO4 , 0.1% Triton X-100, 0.5 μl of 8 U Bst DNA polymerase (New England Biolabs, USA), 40 U of reverse transcriptase (Invitrogen), and 5 μl of target RNA.
Techniques: RT Lamp Assay, Agarose Gel Electrophoresis, Marker, Concentration Assay