Method for predicting calcium antagonist medicine effect and application thereof
A technology of calcium antagonists and drug efficacy, applied in biochemical equipment and methods, medical preparations containing active ingredients, pharmaceutical formulas, etc., can solve problems such as delayed treatment timing, economic losses for patients, and inability to improve drug efficacy and side effects
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Embodiment 1
[0058] Example 1: Determining the Glu429Ala(AC) polymorphism site of the MTHFR gene and predicting the antihypertensive effect of the calcium antagonist class antihypertensive drug nifedipine
[0059] (1) Determination of the genotype of the Glu429Ala(AC) polymorphism site of the MTHFR gene:
[0060] (1) Extract the genomic DNA of the host cell:
[0061] (a) Add 30ml of erythrocyte lysate to the whole blood, shake slowly, and let stand at room temperature for 10 minutes. During this period, shake several times to completely lyse the erythrocytes;
[0062] (b) Centrifuge at 4°C at 2000 rpm for 10 minutes, remove the supernatant, break up the precipitated leukocytes on a rotary shaker, add 40ul of protease and 50ul of RNase, shake well, add 15ml of leukocyte lysate, Mix well in a 37°C water bath for 20 minutes, take it out, and put it in cold water;
[0063] (c) Add 4ml of cold protein precipitation solution, mix well and place in -20°C refrigerator for 5 minutes, take it out ...
Embodiment 2
[0094] Example 2: Determination of the Ala222Val polymorphism site of the MTHFR gene and prediction of the antihypertensive effect of the calcium antagonist antihypertensive drug nifedipine
[0095] (1) Determination of the genotype of the Ala222Val polymorphism site of the MTHFR gene:
[0096] (1) On the standard operating method and operating procedures, adopt the same conventional method as described in Example 1 to extract the genomic DNA of the host cell
[0097] (2) Detection of the MTHFRAla222Val polymorphic site by PCR and restriction enzyme fragment length polymorphism analysis method (PCR-RFLP):
[0098] PCR-specific primers were designed according to the MTHFR Ala222Val gene sequence, including PCR forward primers and PCR reverse primers, and conventional PCR amplification was performed according to the following conditions.
[0099] Primer sequence:
[0100] Sense: 5'-CAAAGG CCA CCC CGAAGC-3'
[0101] Antisense: 5'-AGG ACG GTG CGG TGA GAG TG-3'
[0102] PCR rea...
Embodiment 3
[0120] Example 3: Determination of the MTRR His595Tyr (CT) polymorphism site and prediction of the antihypertensive effect of the calcium antagonist class antihypertensive drug nifedipine
[0121] (1) Determination of the genotype of the His595Tyr (CT) polymorphic site of the MTRR gene:
[0122] (1) Extract the genomic DNA of the host cell:
[0123] On the basis of standard operating procedures, the genomic DNA in salivary cells was extracted by salt precipitation. Store DNA at -20°C.
[0124] (2) The genotype of MTRR His595Tyr (C1894T) polymorphism was detected by TaqMan method.
[0125] (a) Amplify the MTRR His595Tyr gene polymorphic site and its flanking sequences with a PCR instrument, containing 10ng of genomic DNA in a 5ul PCR reaction system, and PCR Master Mix NoAmpErase UNG of 2.5ul TaqMan 2X Universal (composition includes: AmpliTaq Gold DNA Polymerase, dNTPs with Dutp, Passive Reference, optimized buffer), and 0.72uM forward primer, 0.72uM reverse primer and two ...
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