Isothermal Amplification: Enables running amplification reactions using less complex and lower cost instruments
Lyophilization-ready: All kit components are lyophilization compatible, thus avoiding redesign of assays to remove components known to inhibit lyophilization
Fully Optimizable: Non-master mix kit format gives you complete control of reaction formulation to maximize assay performance
Matched Performance to LavaLAMP™ RNA Master Mix: Assists current users of the master mix product by streamlining additional assay optimization with this kit.
Customizable: Custom volumes, dispensing and master mix formulation available to match your specific requirements – contact us
Like the related, LavaLAMP™ RNA Master Mix, this component kit is built around a proprietary DNA polymerase with strand displacement activity that works on both RNA and DNA templates. With these unique features, it is possible to perform RNA LAMP (loop-mediate isothermal amplification starting with RNA targets/templates) using a single DNA polymerase. This enzyme first copies the RNA into cDNA, and then loop mediated isothermal amplification takes over to amplify the initial RNA target sequence present in the new cDNA making amplified and detectable target-specific, dsDNA.
The LavaLAMP™ RNA Component Kit is a deconstruction of the LavaLAMP™ RNA Master Mix, and as such, when reactions are setup using the initial Experimental reaction conditions outlined in the Component Kit User Manual, the composition of that reaction matches the composition of the recommended initial Master Mix reaction outline in its User Manual. The individual reagent format of this kit, as opposed to the master mix format, enables complete control of each LAMP assay allowing you to carefully identify the best reaction conditions for your specific target and primer set.
To aid in assay development and future lyophilization of LavaLAMP RNA-enzyme based reactions, all components in this kit are compatible with lyophilization. This feature means that reactions will not have to be reformulated or reoptimized after removal of components that prohibit lyophilization such as glycerol and betaine.
For more information on loop-mediated isothermal amplification, please review the following recorded webinar, “Loop-Mediated Isothermal Amplification (LAMP): Assay Development Challenges and Solutions.”
Equivalent Performance Between the LavaLAMP™ RNA Master Mix and RNA Component Kit
Figure 1. Comparing the performance of the LavaLAMP™ RNA Component Kit and the LavaLAMP™ RNA Master Mix. LAMP reactions (6 replicates per test) were set up using the indicated kits and targets (A, MS2 and B, Zika) with the same concentrations of Target-Specific Primer Mixes (1X) and the same target input amounts. The LavaLAMP RNA Component Kit reactions were formulated to 1X LavaLAMP RNA Buffer, 5 mM MgSO4 and 0.8 mM dNTPs (each) to match the final formulations of the LavaLAMP RNA Master Mix reactions. Green Fluorescent Dye was included in all reactions. Reactions were run on a CFX96 Thermal Cycler (Bio-Rad) at 68°C for both the MS2 and Zika targets and fluorescence was measured during the course of the 60-minute reactions to determine TTR. NTC denotes No Target Control.
Faster, More Sensitive Detection with LavaLAMP™ RNA Component Kit
Figure 2. Performance comparison of the LavaLAMP™ RNA Component Kit vs. competitor kits. (A) RNA LAMP reactions were set up using the indicated kits according to manufacturer’s recommendations. Target MS2 RNA at varying input amounts, MS2 target RNA LAMP primers, and Green Fluorescent Dye (LavaLAMP Kit) were included in all reactions. Reactions were run on a CFX96 Thermal Cycler (Bio-Rad) at the following temperatures: LavaLAMP; 68°C; other polymerases at the recommended 65°C and fluorescence was measured over 60 minutes to determine the TTR. (B) Delta TTRs were calculated by subtracting the low target level TTR from the NTC TTR for each polymerase tested. Higher positive values indicate better signal to noise, and hence better performance. NTC denotes No Target Control.
Highly Consistent Assay Performance Between Operators
Figure 3. Analysis of operator to operator variability. On the same day, six replicate RNA LAMP reactions were set up per condition per by three different operators using the indicated amounts of MS2 target RNA, MS2 target RNA LAMP primers, and Green Fluorescent Dye. Reactions were run on a CFX96 Thermal Cycler (Bio-Rad) at 68°C and amplification was monitored in real-time. Results were analyzed to determine the TTR and averaged. NTC denotes No Target Control.
Consistent Day to Day Assay Performance
Figure 4. Analysis of day to day assay variability. On three different days, a single operator set up six replicate LAMP reactions per condition tested using the indicated amounts of MS2 target RNA, MS2 target RNA LAMP primers, and Green Fluorescent Dye. On each day the reactions were run on a CFX96 Thermal Cycler (Bio-Rad) at 68°C, amplification was monitored in real-time. The results were analyzed to determine the TTR, and the replicates were averaged and presented. NTC denotes No Target Control.
|SDS||SDS_30096_LavaLAMP RNA Component Kit.pdf|
|SDS||SDS_30097_LavaLAMP RNA Component Kit with Green Fluorescent Dye.pdf|