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A Copper Ion Cutting General-purpose Partition Ultrafast Amplification Visual Sensor
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A technology of copper ion detection and substrate chain, applied in the field of biosensors, can solve the problems of cumbersome sample processing, expensive, and not universally applicable
Active Publication Date: 2020-10-02
CHINA AGRI UNIV
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[0003] Common detection methods for copper ions mainly include atomic spectroscopy, including atomic absorption spectroscopy, inductively coupled plasma emission spectrometry and fluorescence spectroscopy, visible spectrophotometry, flow injection chemiluminescence and differential potential stripping, etc., but these The detection methods all require expensive instruments and professionally trained operators, and the sample pretreatment is also cumbersome and not universally applicable
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Embodiment 1
[0099] Example 1 Establishment of a Copper Ion Cutting Type Universal Interval Ultrafast Amplification Visual Sensor
[0100] 1. Experimental materials
[0101] SYBR Gold nucleic acid dye, nucleic acid molecular weight standard ultra-low range DNA ladder, dNTP, ExTaq DNA polymerase, 10×Taq buffer, hemin, copper chloride, ascorbic acid, 2,2-azino-bis(3-ethane) Base-benzothiazole-6-sulfonic acid) diamine salt (ABTS), H 2 o 2 , 4-Hydroxyethylpiperazineethanesulfonic acid (HEPES), MES, LiCl, Tris, potassium chloride, sodium chloride, magnesium chloride, disodium edetate, sulfuric acid, tetramethylbenzidine, urea, all Purchased from Thermo Scientific Life Technologies. All experimental water was from Milli-Q pure water system.
[0102] The sequence design is as follows (SEQ ID NO:1-7):
[0103]
[0104]
[0105] Note: The ribozyme cuts the target product and is complementary to the 3' end sequence of the amplified template; the underline is to distinguish the size of the...
Embodiment 2
[0127] The specificity investigation of embodiment 2 sensor
[0128] According to the biosensor constructed in Example 1, 10nM Cu 2+ , 10 μM Mg 2+ 、Cd 2+ , Ca 2+ , Zn 2+ 、Cr 3 + , Pb 2+ , Fe 2+ Added to the system for detection, the results show that the established Cu 2+ The biosensor has good specificity ( Figure 5 ).
Embodiment 3
[0129] Embodiment 3 standard addition experiment
[0130] Get high-purity water and detect with the biosensor constructed in embodiment 1, Cu 2+ No detection was carried out, and the standard addition experiment was carried out on it, and the results obtained by continuous measurement are shown in Table 1.
[0131] Table 1 Cu 2+ Spike recovery test results
[0132]
[0133]
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Abstract
The invention provides a copper ion cutting type universal partition superfast amplification visual sensor. The copper ion cutting type universal partition superfast amplification visual sensor comprises: (1) a cutting system; (2) an sPCr amplification system; (3) an HCR system; and (4) a detection system containing ABTS developing liquid. A primer, a template and a probe are designed cleverly, sothat the template is subjected to superfast amplification under the existence of the copper ion; and the amplification product promotes HCR reaction, so that the product forms a G-quadruplex under the proper environment. The peroxidase-like activity of the G-quadruplex is further utilized to develop, so that the problem that the traditional PCR product is difficult in visual detection is solved and rapid and visual detection of the copper ion is realized. More than that, the sensor provided by the invention has the characteristics of high specificity and high sensitivity; and the detection result is more objective and accurate.
Description
technical field [0001] The invention relates to the technical field of biosensors, in particular to a copper ion-cutting general-purpose barrier ultrafast amplification visualization sensor. Background technique [0002] Copper is a transition element, chemical symbol Cu, atomic number 29, atomic weight 63.546, belonging to group IB. Pure copper is a soft metal, its surface is reddish orange with metallic luster when it is just cut, and its single substance is purplish red. Good ductility, high thermal conductivity and electrical conductivity. The amount of copper in the normal human body is 100-200 mg. In the human body, copper ions are mainly used as catalytic cofactors or structures for many enzymes and proteins, and are widely involved in many important metabolisms in the body. The process affects the generation of human blood, the formation of connective tissue, the central nervous system, the metabolism of cholesterol and glucose, the heart function and the immune sys...
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