Primer set, application, product and method for detecting SNP sites related to drug metabolism in children
A primer set and children's technology, applied in biochemical equipment and methods, recombinant DNA technology, microbial measurement/inspection, etc., can solve the problems of long detection cycle, high cost, high technical requirement threshold, etc., and achieve low detection cost and accurate The effect of high reliability and shortened detection cycle
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Embodiment 1
[0099] The design and optimization of embodiment 1 primer, and the establishment of reaction system
[0100] Through the primer design software on the MassARRAY website, adjust the relevant parameters, complete the PCR and UEP primer design of 57 sites, export the designed primers and each parameter file, and synthesize the primers. Prepare amplification primer MIX and extension primer MIX according to the primer configuration table, and fine-tune the extension primer MIX until it meets the requirements. Primer testing and optimization were then performed. Specific steps are as follows:
[0101] (1) Dilute the genomic DNA sample to 10ng / μL, and prepare the PCR reaction MIX according to the table below (the following is the amount of a single sample)
[0102] Table 3 PCR reaction system
[0103]
[0104] Seal the film, mix with vortex for 30 seconds, centrifuge at 4000rpm for 1 minute, and put the plate on the PCR machine for the following thermal cycle:
[0105]
[0...
Embodiment 2
[0147] Verification of embodiment 2 reaction system
[0148] During the system optimization process of this example, 2 to 3 samples of each detection site were verified by Sanger sequencing, and the comparison results were all consistent, confirming that the detection results of this example are accurate. In this example, after confirming the optimal reaction system, a series of verification experiments were carried out, including accuracy, precision, personnel comparison and comparison experiments between different batches of primers. The specific verification scheme is as follows:
[0149] (1) Accuracy experimental verification plan: select one sample from each of the 57 sites for Sanger sequencing, and compare the results of Sanger sequencing and MassARRAY. If the consistency is greater than 95%, the verification is passed.
[0150] (2) Precision experiment verification plan: pick 3 peripheral blood samples and 3 corresponding oral swab samples, repeat 3 times for each sam...
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