TABLE 2.

Direct raw sewage reverse-transcriptase quantitative loop-mediated isothermal amplification (RT qLAMP) reaction mix components

Reaction mix component	Volume
WarmStart master mix (1700 or 2019)	12.5 μL, includes dNTPs at 1.4 mM; RTx, 8 mM MgSO4
10× primers	2.5 μL (1.6 μM FIP/BIP, 0.2 μM F3/B3, 0.4 μM loop F/B)
NEB green fluorescent dye with E1700 kit	0.5 μL (@0.5 μM)
Nuclease-free H2O	4.5–0 μL (adjust w/ template to 25 μL total rxn volume)
Template: control or raw sewage	5–9.5 μL
Total reaction mix	25 μL

Abbreviations: RTx, reverse transcriptase; BIP, backward inner primer; dNTP, deoxynucleoside triphosphate; F/B, forward and backward loop; FIP, forward inner primer; rxn, reaction.