Micro-droplet preparation method applicable to micro-droplet digital PCR (Polymerase Chain Reaction)

Abstract

The invention discloses a micro-droplet preparation method applicable to micro-droplet digital PCR (Polymerase Chain Reaction). The method comprises the following steps: preparing a PCR aqueous phase, wherein the PCR aqueous phase comprises an upstream primer, a downstream primer, a PCR template, a PCRmix and double distilled water; sequentially and uniformly mixing the medicines as a reaction aqueous phase standby; sequentially and uniformly mixing the upstream primer, the downstream primer, the PCR template and the PCRmix, dropwise adding the double distilled water, and then taking a solution system as the reaction aqueous phase standby; preparing a PCR oil phase, and sequentially and uniformly mixing glycerin, Trition X-100, Tween-80 and nonyl phenyl ether IgelCO520; adding 1vol of the PCR aqueous phase into 2vol of the PCR oil phase, and performing vortex stirring and reaction to obtain water-in-oil micro-droplets. The preparation method provided by the invention has the advantages that the water-in-oil reaction micro-droplets which are uniform in granularity and stable in thermodynamic property can be conveniently prepared, and the operation of a micro-droplet digital PCR experiment is simplified.

Description

technical field [0001] The invention relates to the field of molecular diagnosis, and is mainly applied to microdroplet preparation in digital microdroplet PCR or other PCRs. Background technique [0002] The particle size of the nanospheres prepared by the traditional inverse microemulsion method is generally about 10-100 nanometers, not only small in size, but also hollow. It is mostly used for chemical modification on its surface. The present invention provides the establishment of a new method. On the basis of the improved inverse microemulsion method, water-in-oil immiscible properties are used to form monodisperse water-in-oil droplets one by one, with a particle size of about 10- Between 100 μm, it can be used as a micro-container for many biochemical reactions due to the aqueous solution contained in it. [0003] The traditional PCR method is carried out in PCR aqueous solution, because homologous base sequences will be complementary between templates and primers, ...

AI Technical Summary

Problems solved by technology

[0003] The traditional PCR method is carried out in PCR aqueous solution, because homologous base sequences will be complementary between templates and primers, resulting in the amplification of non-specific bands in PCR, which often increases the generation of false positives

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