Chemiluminescence sensing method based on dsDNA-SYBR Green I light-sensitive catalysis
A chemiluminescence and sensing method technology, applied in the field of biological sensing, can solve the problem of high background, achieve the effects of improving sensitivity, high sensitivity and reducing detection cost
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Embodiment 1
[0016] Take a certain volume of BRCA1 gene (5'-GAG CAT ACA TAG GGTTTCTCTTGGTTT-3'), 15 μL of 200 nM BRCA1 complementary chain (CS-BRCA1: 5'-AAA CCA AGA GAAACC CTA TGTATG CTC-3') in Add a certain volume of NaH to a multiwell plate with a volume of 300 μL 2 PO 4 After incubating the buffer solution for 60 min, add 2.4 μL of SG dye at a concentration of 250× and incubate for 15 min; then add 15 μL of K at a concentration of 1 mM 4 [Fe(CN) 6 ] solution, placed under a blue LED light (3W) for 30 min. Finally, 100 μL of the sample after illumination was added to 100 μL of 2.0 mM luminol (pH = 11.5). At room temperature, the chemiluminescence intensity was measured with a chemiluminescence instrument, and the concentration of the BRCA1 gene was detected based on the obtained signal intensity.
Embodiment 2
[0018] Take a certain volume of BRCA2 gene (5'-AAA GGGCTTCTG ATT-3'), 15 μL of BRCA2 complementary strand (CS-BRCA2: 5'-AAT CAG AAG CCC TTT-3') at a concentration of 200 nM in a volume of 300 μL Add a certain volume of NaH to the perforated plate 2 PO 4 After incubating the buffer solution for 60 min, add 2.4 μL of SG dye at a concentration of 250 × and incubate for 15 min; then add 15 μL of K at a concentration of 1 mM 4 [Fe(CN) 6 ] solution, placed under a blue LED light (3W) for 30 min. Finally, 100 μL of the sample after illumination was taken and added with 100 μL of 2.0 mM luminol (pH = 11.5). At room temperature, the chemiluminescence intensity was measured with a chemiluminescence instrument, and the concentration of the BRCA2 gene was detected based on the obtained signal intensity.
Embodiment 3
[0020] Take a certain volume of p53 gene (5'-TTC CTC TGTGCGCCGGTCTCTCCT-3'), 15 μL of p53 complementary strand (CS-p53: 5'-AGG AGA GACCGGCGCACA GAG GAA-3') at a concentration of 200 nM in a volume of 300 μL Add a certain volume of NaH to the perforated plate 2 PO 4 After incubating the buffer solution for 60 min, add 2.4 μL of SG dye at a concentration of 250 × and incubate for 15 min; then add 15 μL of K at a concentration of 1 mM 4 [Fe(CN) 6 ] solution, placed under a blue LED light (3W) for 30 min. Finally, 100 μL of the sample after illumination was taken and added with 100 μL of 2.0 mM luminol (pH = 11.5). At room temperature, the chemiluminescence intensity was measured with a chemiluminescence instrument, and the concentration of the p53 gene was detected based on the obtained signal intensity.
[0021] The detection limits of BRCA1, BRCA2, and p53 genes detected by Examples 1, 2, and 3 were 3.0 pM, 6.0 pM, and 5.0 pM, respectively. This system can be applied to the...
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