Kit for accurately detecting polymorphism of ALDH2 (Aldehyde Dehydrogenase 2) gene
A kit and site polymorphism technology, applied in the field of biomedicine, can solve problems such as complex operation process, wrong diagnosis results, long detection cycle, etc., and achieve the effects of high sensitivity, simple operation, and convenient sampling
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Embodiment 1
[0029] Example 1: Determination of genotyping criteria
[0030] Samples of accurate genotypes obtained by Sanger sequencing were collected, including 100 cases of wild type, mutant type and heterozygous type, and then detected by the kit of the present invention.
[0031] 1. Multiplex PCR amplification reaction
[0032] 1.1 Reagent preparation: Return the PCR reaction solution, primer mixture, saliva collector, saliva preservation solution, quality control and sterilized ultrapure water in the kit to room temperature.
[0033] 1.2 Construction of PCR amplification system
[0034] The kit of the present invention uses a 25 μL reaction system, and the above reagents are prepared according to the proportions described in Table 2.
[0035] Mix, dispense into PCR reaction tubes, add the samples to be tested on the machine for detection.
[0036] Table 2 Multiplex PCR amplification system
[0037] Reagent
volume
PCR reaction solution
10 μL
primer m...
Embodiment 2
[0042] Example 2: Peripheral blood sample detection
[0043] Take 30 cases of fresh whole blood treated with EDTA, and use the kit of the present invention to directly detect. The multiplex PCR amplification curve was analyzed, and the results were compared with the Sanger sequencing method, and the results are shown in Table 4. The results show that the coincidence rate of the genotyping results of the kit of the present invention and the results of the Sanger sequencing method is 100%.
[0044] Table 4 Genotyping accuracy of whole blood samples
[0045] sample number
Embodiment 3
[0046] Example 3: Saliva sample detection
[0047] Use the saliva collector included in the kit to collect the saliva of the test subject, put it in the saliva preservation solution and mix it upside down, take 1 μL as the PCR amplification template, and mix evenly according to the multiplex PCR amplification system shown in Table 1, and the PCR amplification system shown in Table 2 Amplification Conditions PCR amplification was performed. The test results of 29 samples to be tested were compared with the Sanger sequencing method, and the results are shown in Table 5.
[0048] Table 5 Genotyping accuracy of saliva samples
[0049] sample number
[0050] The results show that the coincidence rate of the genotyping results of the kit of the present invention and the results of the Sanger sequencing method is 100%.
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