EA0201 LAMP Colorimetric Master Mix (Red to Yellow)

LAMP Colorimetric Master Mix (Red to Yellow)

Cat#EA0201

Description

This product is a pH sensitive LAMP reaction mix. The mix is optimised with buffer, colour dye, Bst DNA polymerase, Mg²⁺ and dNTP in a special Low Salt buffer.

During the LAMP reaction, Bst DNA polymerase plays the role of polymerisation, changing the number of protons and thus the pH value, turning the red reaction solution into yellow, the whole reaction process is fast and the colour change is clear and distinguishable to the naked eye. To use, all that is required is that the primers are mixed and added to the sample, the reaction is kept at a constant temperature, and a positive amplification can be read with the naked eye within 20-40 minutes.

Product Composition

Storage Conditions

Dry ice transportation, please be careful to avoid repeated freeze-thaw. Store at -20 ℃ for 6 months.

Experimental Procedures

1. This product will have a slight sediment after removal at -20 ℃. Thaw and mix at room temperature until the precipitation disappears, and then use after centrifugation.

2. Configure the reaction mixture according to the following components

Mix gently and centrifuge.

* Recommended final primer concentration: 1.6 μM FIP/BIP, 0.4 μM LF/LB, 0.2 μM F3/B3.

** The amount of template to be added should be adjusted within 1-8 µL, and it is recommended to dilute the template with water because the reagent is very sensitive to pH, so avoid using Tris or other buffer systems to dilute the template to avoid the change of pH of the system, which may prevent the positive reaction from generating the discolouration reaction.

3. After placement in the PCR instrument, react at 63 ℃ for 40 minutes (hot cover temperature 105 ℃) and 85 ℃ for 5 minutes.

Notes:

1) The enzymes used in this kit are very sensitive to temperature, and it is strongly recommended to use a PCR instrument for operation; if a water bath is used, it should be heated first to reach the specified temperature before reaction. Too large a temperature difference will result in lower amplification efficiency and prevent the positive reaction from turning red to yellow within the time specified in the instructions.

2) Negative controls: It is recommended to set up a blank (no template) control as well as an NTC (usually water) control.

Results Interpretation:

1. Immediately after the experiment, take out the reaction tube and leave it at room temperature for 5 minutes to observe the result. Observe the results in a well-lit environment (white background is recommended). If the reaction solution is yellow in colour, it will be judged as positive, if the reaction solution is purplish red, it will be judged as negative, and if the reaction solution is orange-yellow, it will be judged as weakly positive.

2. It is forbidden to open the cap of the reaction tube at the end of the experiment to prevent aerosol contamination.

3. The reaction time must be timed accurately, exceeding the reaction time specified in the instructions may result in false positives.

Considerations

1. The operations of reagent preparation and template addition are carried out in different areas to avoid contamination of the environment and subsequent experiments.

2. The reagents should be avoided to be exposed to air for a long time after dispensing.

3. The red-yellow colour change reaction depends on the pH change of the reaction system, please do not use nucleic acid preservation solution containing high concentration of Tris, it is recommended to use water for preservation.

4. 1 fg-10 ng of plasmid DNA and 1pg-100ng of genomic DNA are recommended.

Examples of applications

Legend: amplification of the new crown N gene sensitivity test