Medication guiding gene detection reagent kit of hypertension reducing medicine namely trandolapril
A technology for trandolapril and hypertension, applied in the field of oligonucleotide products, can solve the problems of difficult to meet the needs of fast and accurate, cumbersome operation, limited detection objects, etc.
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Embodiment 1
[0085] Example 1: Primer Design and Synthesis
[0086] The surrounding sequences of the target SNP of this kit are queried in the db_SNP (build 132) and Hapmap (Rel 28, Phase II + III, Aug 10) databases, and these sequences are used to design multiple PCR primers and single base extension primers.
[0087] The corresponding specific PCR primer core sequences (SEQ1a to SEQ5a) and specific extension primer core sequences (SEQ1b to SEQ5b) were designed for five polymorphic sites related to the discrimination of drug types, including rs699, rs2016848, rs11209716, rs4742610, and rs8012552. 5 pairs of PCR primers and 5 extension primers form 3 independent reaction systems: SEQ1a / b to SEQ2a / b form the first reaction system, SEQ3a / b to SEQ4a / b form the second reaction system, and SEQ5a / b forms the The third reaction system. In these 3 independent reaction systems, SEQ1a to SEQ2a, SEQ3a to SEQ4a, and SEQ5a respectively participate in 3 independent multiplex PCR reactions, and SEQ1b to...
Embodiment 2
[0089] Embodiment 2: sample DNA extraction
[0090] A total of 10 DNA samples were collected from ordinary Chinese people, marked as A1-A10. Among them, sample collection, DNA extraction, etc. were collected in accordance with the requirements of the instructions, and human venous blood was collected with EDTA anticoagulant tubes. According to the instructions, the collected blood should not be stored at 2-8°C for more than one week, and at -20°C for no more than one month, and can be transported in a curling box with ice or a foam box with ice. It is recommended to use fresh blood as much as possible. Genomic DNA extraction. Since this kit does not provide human genomic DNA extraction reagents, a commercial nucleic acid extraction kit (such as QIAGEN’s DNeasy Blood and Tissuekit) was used to extract human genomic DNA from 200 μL of whole blood of each patient, and the DNA was extracted using a NanoDrop 2000 ( Thermo Company) quantified, and standardized to 30ng / μL (respecti...
Embodiment 3
[0091] Embodiment three: biological experiment
[0092] Using ABI 9700 type PCR instrument, according to the instructions, check the 5 polymorphic sites that are used to identify the drug type.
[0093] The components used in the kit for PCR, PCR product purification and single base extension are:
[0094] serial number component name main ingredient Specification 1 PCR Primer Mix PCR primers 24μL / tube x1 tube 2 PCR reaction solution Taq enzyme, dNTP 72μL / tube x1 tube 3 Enzyme digestion reaction solution SAP enzyme 48μL / tube x1 tube 4 Extension Primer Mix extension primer 24μL / tube x1 tube 5 Extension reaction solution Single base elongase, ddNTP 24μL / tube x1 tube 6 positive control Human Genomic DNA (30ng / μL) 10μL / tube x1 tube
[0095] The concentration of each primer pair is 500nmol / L.
[0096] According to the manual, the specific operation method is as follows:
[0097] 1. PCR amplification ...
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