Method for detecting target numerator based on nucleic acid aptamer and PCR amplification
A nucleic acid aptamer and target detection technology, which is applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of limited application, achieve the effect of few influencing factors, simple operation and steps, and reliable results
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Embodiment 1
[0032] Example 1: Use Aptamer based PCR to detect trace amounts of cocaine in serum or plasma (small molecule detection) (without magnetic bead enrichment and separation)
[0033] DNA sequences used in this experiment:
[0034] A chain sequence: (39 bases)
[0035] 3-HS-(CH2) 6 -CTG GGT GAA GTA ACT TCC TAA AAG GAA CAG AGG GGGAGT-5
[0036] B chain sequence: (58bases)
[0037] 5-GTGTGAAGCGGCATCTAAAGCGTCTCCCCCCTCAAGACTAGGTCCAGCCCAGGGTGTC-biotin-3
[0038] The part in italics is the hybridization site of the template
[0039] B chain primer:
[0040] S: 5-GTG TGA AGC GGC ATC TAA AGC-3 (21bases)
[0041] A: 5-GAC ACC TCT GGG CTG GAC C-3 (19bases)
[0042] Experimental steps:
[0043] 1. Treatment of gold electrodes
[0044] 1. First drop a drop of separation reagent on the gold electrode, after 15 minutes, wash it off with double distilled water.
[0045] Separation reagent: H 2 o 2 : Concentrated H 2 SO 4 =7:3 (volume ratio)
[0046] 2. Use Al 2 o 3The slurry pol...
example 2
[0089] Example 2: Using Aptamer based PCR to detect thrombin in serum (protein detection)
[0090] DNA sequences used in this experiment:
[0091] A chain sequence:
[0092] 5-HS-(CH2) 6 -ATATAGGTTGGTGTGGTTGG-3
[0093] B chain sequence:
[0094] 5-CAG TGA AGC GGC ATC TAA AGC ACG AGT CTG AGT CCA ACC ACA-3
[0095] Primer for B chain: A: 5-TGT GGT TGG ACT CAG AC TCG-3
[0096] S: 5-GTG TGA AGC GGC ATC TAA AGC-3
[0097] Experimental steps:
[0098] A gold electrode treatment:
[0099] 1 First drop a drop of separation reagent on the gold electrode, after 15 minutes, wash it off with double distilled water.
[0100] Separation reagent: H 2 o 2 : Concentrated H 2 SO 4 =7:3 (volume ratio)
[0101] 2 with Al 2 o 3 The slurry polishes the gold electrodes. When grinding the gold electrode, the force should be uniform, and the gold electrode must be vertical.
[0102] 3 Rinse the gold electrode with double distilled water, and dry the electrode with filter paper and e...
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