CN113005184A - Reagent and method for enhancing real-time fluorescent PCR (polymerase chain reaction) signal - Google Patents
Reagent and method for enhancing real-time fluorescent PCR (polymerase chain reaction) signal Download PDFInfo
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- CN113005184A CN113005184A CN202011513133.0A CN202011513133A CN113005184A CN 113005184 A CN113005184 A CN 113005184A CN 202011513133 A CN202011513133 A CN 202011513133A CN 113005184 A CN113005184 A CN 113005184A
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Abstract
The invention provides a reagent for enhancing a fluorescence PCR signal and application thereof, wherein the reagent comprises an enhancer containing betaine and dimethyl sulfoxide. When detecting a gene with higher GC content near a site to be detected, the reagent and the method provided by the invention can make the genotyping of the gene site in a high GC content area more effective, and specifically, the Ct value is smaller, the signal value is higher, and the non-specific signal is lower.
Description
Technical Field
The present invention relates to the field of biotechnology, and more particularly to reagent compositions and methods for enhancing fluorescent PCR signals.
Background
Real-time fluorescent PCR technology, which monitors the amplification of PCR products by introducing specific probes and monitoring fluorescent signals in real time. Compared with a sequencing method, a gene chip method, an allele specific PCR technology and a high resolution melting curve method, the whole real-time fluorescence PCR process can be completed in a single tube in a closed manner, and the result can be automatically analyzed. At present, the real-time fluorescent PCR method is used for detecting gene polymorphism and microbial pathogens such as viruses, and the like, and the application is more and more extensive.
DNA sequences with high GC content are widely present in the genome and they are particularly prone to secondary structure interference PCR. The real-time fluorescent PCR technology is used for gene polymorphism detection, and primers and probes need to be designed for sites to be detected. Due to the particularity of the detection principle, the site to be detected is generally positioned at one third of the 3' end of the probe when the probe is designed, and the length of an amplification product is generally controlled to be 70bp-150 bp. Because the design is carried out around the site to be detected, the selection space of the primer and the probe sequence is smaller, when the GC content near the site to be detected is higher, firstly, the GC content of the selected primer and probe sequence is inevitably higher, and secondly, a DNA template easily forms a stable hairpin loop secondary structure, so that the nonspecific combination of the primer and probe is caused, and further, the problems of large Ct value, weak fluorescence signal, high nonspecific signal and the like are caused.
Disclosure of Invention
The invention aims to provide a real-time fluorescence PCR enhancer to overcome the problems of high GC content, large Ct value, weak fluorescence signal, high non-specific signal and the like caused by the secondary structure of a hairpin loop and the like caused by high GC content.
In order to realize the purpose of the invention, the technical scheme of the invention is as follows:
providing an enhancer for real-time fluorescent PCR, comprising betaine and dimethyl sulfoxide; the concentration ratio of the betaine to the dimethyl sulfoxide is (4M-4.62M) to (7.69 v/v% -20%) (volume percentage).
In some embodiments, the betaine and dimethyl sulfoxide are present in the enhancer in a volume ratio of 4:1 to 12:1 when the betaine concentration is 5M and the dimethyl sulfoxide concentration is 100%. When the concentrations of betaine and dimethyl sulfoxide are changed, the proportion of each component is properly adjusted according to the concentration of each component by those skilled in the art, and all that is included in the protection scope of the present invention.
In some embodiments of the invention, the enhancer consists of betaine, dimethyl sulfoxide. The concentration of betaine is 5M, the concentration of dimethyl sulfoxide is 100%, and the volume ratio of betaine to dimethyl sulfoxide is 4: 1. The final concentration of the enhancer added into a fluorescent PCR reaction system is 0.2M of betaine and 1% of dimethyl sulfoxide.
In some embodiments of the invention, the enhancer consists of betaine, dimethyl sulfoxide. The concentration of betaine is 5M, the concentration of dimethyl sulfoxide is 100%, and the volume ratio of betaine to dimethyl sulfoxide is 8: 1. The final concentration of the enhancer added into a real-time fluorescent PCR reaction system is 0.6M of betaine and 1.5% of dimethyl sulfoxide.
In some embodiments of the invention, the enhancer consists of betaine, dimethyl sulfoxide. The concentration of betaine is 5M, the concentration of dimethyl sulfoxide is 100%, and the volume ratio of betaine to dimethyl sulfoxide is 12: 1. The final concentration of the enhancer added into a real-time fluorescent PCR reaction system is 1.2M of betaine and 2% of dimethyl sulfoxide.
A method for enhancing a fluorescence PCR signal comprises the steps of configuring a real-time fluorescence PCR reaction system, and adding an enhancer into the real-time fluorescence PCR reaction system, wherein the enhancer comprises betaine and dimethyl sulfoxide.
In some embodiments of the invention, the final concentration of betaine added to the real-time fluorescent PCR reaction system is 0.2M to 1.2M.
In some embodiments of the invention, the final concentration of dimethyl sulfoxide added to the real-time fluorescent PCR reaction system is 1% to 2% (volume percent).
The invention also provides a reaction system of real-time fluorescence PCR, which comprises Taq enzyme, dNTP and PCR buffer solution and is characterized by also comprising an enhancer, wherein the enhancer comprises betaine and dimethyl sulfoxide.
In some embodiments of the invention, the final concentration of betaine added to the real-time fluorescent PCR reaction system is 0.2M to 1.2M.
In some embodiments of the invention, the final concentration of dimethyl sulfoxide added to the real-time fluorescent PCR reaction system is 1% to 2% (volume percent).
The betaine in the enhancer provided by the invention can improve the hydration of guanine and cytosine in a minor groove of DNA in a high GC region, influence the DNA conformation, relieve the secondary structure, prevent DNA polymerase from dissociating from template DNA, and simultaneously can reduce the annealing temperature of a primer with a high Tm value. Dimethyl sulfoxide in the enhancer can directly remove the secondary structure of template DNA, thereby increasing the specific binding of the primer probe and the template and reducing the non-specific binding. The invention can make the gene locus typing in the high GC content area more effective, and concretely, the Ct value is smaller, the signal value is higher, and the non-specific signal is lower.
Drawings
FIG. 1 shows the real-time fluorescent PCR results of example 2.
FIG. 2 shows the real-time fluorescent PCR results of example 3.
Detailed Description
In order to make the technical methods, advantages and objects of the present invention more comprehensible, the present invention is described in detail below with reference to specific embodiments and accompanying drawings. The examples described herein are intended to be illustrative of the invention and are not intended to be limiting.
Example 1 preparation of enhancer
The raw materials of the enhancer comprise betaine and dimethyl sulfoxide, wherein the concentration of the betaine is 5M, the solvent is water, and the concentration of the dimethyl sulfoxide is 100%. The preparation method comprises mixing 5M betaine and dimethyl sulfoxide at a volume ratio of 8: 1.
Example 2 application in high GC content genotyping
1. DNA sequence to be amplified
The GC content of this DNA region was 76%, and the sites to be detected were in brackets.
2. Primer sequences and probe sequences
The primers and probes described in Table 1 were designed for the sites to be detected according to the DNA sequence to be amplified.
Table 1: primer and probe sequences for real-time fluorescent PCR
| Primer and Probe sequences (5 '-3') | GC content (%) | Tm value (. degree.C.) |
| A forward primer: gctgcaggcggcgcagg | 82 | 68 |
| Reverse primer: cgaggcgcacccgcagc | 82 | 69 |
| Wild-type probe: FAM-ggacgtgtgcggccgc-MGB | 81 | 76 |
| Mutant probes: VIC-ggacgtgcgcggccg-MGB | 86 | 75 |
3. Real-time fluorescent PCR reaction system and program
A real-time fluorescent PCR reaction system was prepared as shown in Table 2.
TABLE 2
| Components | Volume per reaction system |
| Taq enzyme (6U/. mu.L) | 0.5μL |
| dNTP(10mM) | 0.4μL |
| PCR buffer (2X) | 10μL |
| Forward primer (100. mu.M) | 0.1μL |
| Reverse primer (100. mu.M) | 0.1μL |
| Wild type probe (100. mu.M) | 0.05μL |
| Mutant probe (100. mu.M) | 0.05μL |
| Form panel | 2.0μL |
Preparing an enhancer mother solution: the enhancer comprises betaine and dimethyl sulfoxide, wherein the betaine concentration is 5M, the solvent is water, and the dimethyl sulfoxide concentration is 100%.
Experimental group 1: an enhancer was added to the PCR reaction system of Table 2 so that the final concentration of betaine was 0.2M and the final concentration of dimethyl sulfoxide was 1% (by volume), and the mixture was made up to 20. mu.L with sterilized purified water.
Experimental group 2: an enhancer was added to the PCR reaction system of Table 2 so that the final concentration of betaine was 0.6M and the final concentration of dimethyl sulfoxide was 1.5% (by volume), and the mixture was made up to 20. mu.L with sterilized purified water.
Experimental group 3: an enhancer was added to the PCR reaction system of Table 2 so that the final concentration of betaine was 1.2M and the final concentration of dimethyl sulfoxide was 2% (by volume), and the mixture was made up to 20. mu.L with sterilized purified water.
Control group 1: betaine and dimethyl sulfoxide were not added to the PCR reaction system of Table 2, and sterilized purified water was used alone to make up to 20. mu.L.
Control group 2: betaine was added to the PCR reaction system of Table 2 at a final concentration of 0.6M, and dimethyl sulfoxide was not added, and the amount of the mixture was made up to 20. mu.L with sterilized purified water.
Control group 3: dimethyl sulfoxide was added to the PCR reaction system of Table 2 to a final concentration of 1.5% (volume percent) and betaine was not added and made up to 20. mu.L with sterilized purified water.
The specific addition amounts of the experimental group and the control group are shown in Table 3.
TABLE 3
The PCR amplification procedure was: 5min at 95 ℃; at 95 ℃ for 15s, at 60 ℃ for 30s, and for 40 cycles. FAM and VIC channels are selected. After the reaction is finished, the software of the instrument is used for adjusting the starting value, the ending value and the threshold value of the base line to carry out analysis, the Ct value is recorded, and the result is interpreted.
The typing result judging method comprises the following steps:
4. results of the experiment
The results of the experiment are shown in table 4 and fig. 1.
From the experimental results, it can be seen that when only betaine is added to the reaction system, the Ct value can be reduced, the signal can be enhanced (increased Δ Rn), but a strong non-specific signal (the Ct value of the VIC channel is less than or equal to 35) is caused to cause typing errors. When only dimethyl sulfoxide is added to the reaction system, the nonspecific signal is low, but the Ct value is large, the signal is reduced, and the risk of non-typing exists.
Betaine and dimethyl sulfoxide are added into the three experimental groups, so that the Ct value is reduced, the signal is enhanced, the increase of non-specific signals is not caused, the typing effect is greatly improved, and the typing accuracy is improved.
The real-time fluorescent PCR signal can be enhanced by adding an enhancer containing 0.2M-1.2M of betaine and 1% -2% of dimethyl sulfoxide into the reaction system.
Table 4:
| experimental group 1 | |
Experimental group 3 | Control group 1 | |
Control group 3 | |
| Ct(FAM) | 23.38 | 22.25 | 22.89 | 25.17 | 22.61 | 25.82 |
| Ct(VIC) | 36.86 | 38.14 | Undet. | 35.38 | 29.9 | Undet. |
| ΔRn | 1.59 | 1.51 | 1.28 | 1.06 | 1.75 | 0.88 |
| Genotype interpretation | T/T | T/T | T/T | T/T | T/C | T/T |
Example 3 reaction System stability test
1. DNA sequence to be amplified
The GC content of this DNA region was 76%. The sites to be detected are in square brackets.
2. Primer sequences and probe sequences
The primers and probes of Table 7 were designed for the sites to be detected, according to the DNA sequence to be amplified.
Table 7: real-time fluorescent PCR primer and probe sequence
3. Real-time fluorescent PCR reaction system and program
An enhancer comprising betaine and dimethyl sulfoxide was added to the PCR reaction system, the final concentration of betaine was 0.6M and the final concentration of dimethyl sulfoxide was 1.5% (volume percent). See table 5 for details.
TABLE 5
| Components | Volume per reaction system |
| Taq enzyme (6U/. mu.L) | 0.5μL |
| dNTP(10mM) | 0.4μL |
| PCR buffer (2X) | 10μL |
| Forward primer (100. mu.M) | 0.1μL |
| Reverse primer (100. mu.M) | 0.1μL |
| Wild type probe (100. mu.M) | 0.05μL |
| Mutant probe (100. mu.M) | 0.05μL |
| Betaine (5M) | 2.4μL |
| Dimethyl sulfoxide (100%) | 0.3μL |
| Form panel | 2.0μL |
| Purified water | Make up to 20. mu.L |
After the reaction system is prepared, the reaction system is put into a constant-temperature incubator at 37 ℃ for 3, 5 and 7 days for accelerated treatment, and the detection on the computer is completed.
The real-time fluorescent PCR amplification procedure is as follows: 5min at 95 ℃; at 95 ℃ for 15s, at 60 ℃ for 30s, and for 40 cycles. FAM and VIC channels are selected. After the reaction is finished, the software of the instrument is used for adjusting the starting value, the ending value and the threshold value of the base line to carry out analysis, the Ct value is recorded, and the result is interpreted.
4. Results of the experiment
TABLE 6
| |
3 days | 5 |
7 days | |
| Ct(FAM) | Undet. | Undet. | Undet. | Undet. |
| Ct(VIC) | 24.99 | 25.32 | 25.33 | 25.95 |
| Genotype interpretation | C/C | C/C | C/C | C/C |
The results of the experiment are shown in table 6 and fig. 2. The reaction solution added with the reinforcing agent is placed in a constant temperature incubator at 37 ℃ for 3, 5 and 7 days, and compared with the reaction solution without treatment, the Ct value is not obviously increased, the typing is accurate, and the reaction solution added with the reinforcing agent has better stability.
Sequence listing
<120> a reagent and method for enhancing real-time fluorescent PCR signal
<160> 10
<170> SIPOSequenceListing 1.0
<210> 1
<211> 179
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 1
ccccggtggc ggaggagacg cgggcacggc tgtccaagga gctgcaggcg gcgcaggccc 60
ggctgggcgc ggacatggag gacgtgcgcg gccgcctggt gcagtaccgc ggcgaggtgc 120
aggccatgct cggccagagc accgaggagc tgcgggtgcg cctcgcctcc cacctgcgc 179
<210> 2
<211> 17
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 2
gctgcaggcg gcgcagg 17
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<211> 17
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 3
cgaggcgcac ccgcagc 17
<210> 4
<211> 16
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 4
<210> 5
<211> 15
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 5
ggacgtgcgc ggccg 15
<210> 6
<211> 179
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 6
ccccggtggc ggaggagacg cgggcacggc tgtccaagga gctgcaggcg gcgcaggccc 60
ggctgggcgc ggacatggag gacgtgtgcg gccgcctggt gcagtaccgc ggcgaggtgc 120
aggccatgct cggccagagc accgaggagc tgcgggtgcg cctcgcctcc cacctgcgc 179
<210> 7
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<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 7
gctgcaggcg gcgcagg 17
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<212> DNA
<213> Artificial Sequence (Artificial Sequence)
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cgaggcgcac ccgcagc 17
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<212> DNA
<213> Artificial Sequence (Artificial Sequence)
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<210> 10
<211> 15
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 10
ggacgtgcgc ggccg 15
Claims (10)
1. An agent for enhancing a fluorescent PCR signal, comprising an enhancer, wherein the enhancer comprises betaine and dimethyl sulfoxide.
2. The enhancer as claimed in claim 1, wherein the concentration of betaine in the enhancer is 5M.
3. The enhancer of claim 1, wherein the concentration of dimethyl sulfoxide in the enhancer is 100%.
4. The enhancer of claim 1 wherein the betaine and the dimethyl sulfoxide in the enhancer are separately packaged.
5. A method for enhancing a fluorescent PCR signal comprises the step of configuring a real-time fluorescent PCR reaction system, wherein an enhancer is added into the real-time fluorescent PCR reaction system, and the enhancer comprises betaine and dimethyl sulfoxide.
6. The method of claim 5, wherein the final concentration of betaine added to the real-time fluorescent PCR reaction system is 0.2M to 1.2M.
7. The method of claim 5, wherein the final concentration of dimethyl sulfoxide added to the real-time fluorescent PCR reaction system is 1 to 2% (volume percent).
8. A reaction system of real-time fluorescence PCR comprises Taq enzyme, dNTP and PCR buffer solution, and is characterized by further comprising an enhancer, wherein the enhancer comprises betaine and dimethyl sulfoxide.
9. The reaction system of claim 8, wherein betaine is added to the real-time fluorescent PCR reaction system at a final concentration of 0.2M to 1.2M.
10. The reaction system of claim 8, wherein the final concentration of dimethyl sulfoxide added to the real-time fluorescent PCR reaction system is 1-2% (volume percentage).
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102465120A (en) * | 2010-11-10 | 2012-05-23 | 深圳华大基因科技有限公司 | Fluorescence quantitative PCR reaction solution and fluorescence quantitative PCR method |
| CN106498060A (en) * | 2016-11-03 | 2017-03-15 | 江苏然科生物技术有限公司 | A kind of fluorescence quantitative PCR reaction solution and method |
| US20180187251A1 (en) * | 2014-06-02 | 2018-07-05 | Illumina Cambridge Limited | Methods of reducing density-dependent gc bias in amplification |
| CN110804652A (en) * | 2019-12-03 | 2020-02-18 | 郑州安图生物工程股份有限公司 | Additive, kit and reaction method for rapid detection of real-time quantitative PCR of DNA |
| CN111876473A (en) * | 2020-08-07 | 2020-11-03 | 福州大学 | Buffer solution suitable for TTH DNA enzyme amplification in fluorescent quantitative PCR |
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- 2020-12-20 CN CN202011513133.0A patent/CN113005184A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102465120A (en) * | 2010-11-10 | 2012-05-23 | 深圳华大基因科技有限公司 | Fluorescence quantitative PCR reaction solution and fluorescence quantitative PCR method |
| US20180187251A1 (en) * | 2014-06-02 | 2018-07-05 | Illumina Cambridge Limited | Methods of reducing density-dependent gc bias in amplification |
| CN106498060A (en) * | 2016-11-03 | 2017-03-15 | 江苏然科生物技术有限公司 | A kind of fluorescence quantitative PCR reaction solution and method |
| CN110804652A (en) * | 2019-12-03 | 2020-02-18 | 郑州安图生物工程股份有限公司 | Additive, kit and reaction method for rapid detection of real-time quantitative PCR of DNA |
| CN111876473A (en) * | 2020-08-07 | 2020-11-03 | 福州大学 | Buffer solution suitable for TTH DNA enzyme amplification in fluorescent quantitative PCR |
Non-Patent Citations (2)
| Title |
|---|
| 李长贵等: "应用复合增强剂扩增人巨细胞病毒pp65全基因", 《中国生物工程杂志》 * |
| 王同同等: "甜菜碱改善水稻高GC含量DNA序列的PCR扩增", 《生物技术通报》 * |
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