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Nucleic acid release liquid, kit and nucleic acid release method

A nucleic acid release and kit technology, which is used in biochemical equipment and methods, microbial determination/inspection, DNA preparation, etc., can solve the problems of long waiting time, long incubation, precipitation, long and complicated operation steps, etc., to reduce work Volume and time-consuming effect

Pending Publication Date: 2022-06-28
生工生物工程(上海)股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods can usually obtain DNA with high purity and less impurities. However, these methods require long incubation and precipitation, and there are problems such as lengthy and complicated operation steps and long waiting time.

Method used

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  • Nucleic acid release liquid, kit and nucleic acid release method
  • Nucleic acid release liquid, kit and nucleic acid release method
  • Nucleic acid release liquid, kit and nucleic acid release method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] This embodiment provides a nucleic acid release solution and release method (or use method) for blood direct expansion probe method qPCR.

[0051] The nucleic acid release solution contains: 50 mM Tris-HCl (pH 8.0), 35 mM potassium hydroxide, 200 mM ammonium chloride, 0.05% Tween-20 (volume percent concentration).

[0052] Instructions:

[0053] S1. Select a clean 200 μl EP tube, and add 50 μl of nucleic acid release solution to the EP tube.

[0054] S2. Take the same volume of blood sample as the nucleic acid release solution (the source of blood used in the present invention is our company), add it to the above EP tube, vortex to mix well, and centrifuge briefly.

[0055] S3. The above reaction solution is placed in a PCR apparatus, and after reacting at 95° C. for 5 minutes, it is placed in a centrifuge for 2 minutes, and the supernatant can be directly used as a qPCR reaction template.

[0056] Further, in step S3, the centrifugal rotation speed of the centrifuge ...

Embodiment 2

[0058] This embodiment provides a nucleic acid release solution and release method (or use method) for blood direct expansion probe method qPCR.

[0059] The nucleic acid release solution contains: 50 mM Tris-HCl (pH 8.0), 35 mM potassium hydroxide, 200 mM ammonium chloride, 0.2% Tween-20 (volume percent concentration).

[0060] Instructions:

[0061] S1. Select a clean 200 μl EP tube, and add 50 μl of nucleic acid release solution to the EP tube.

[0062] S2. Take the same volume of blood sample as the nucleic acid release solution and add it to the above EP tube, vortex to mix well, and centrifuge briefly.

[0063] S3. The above reaction solution is placed in a PCR apparatus, and after reacting at 95° C. for 5 minutes, it is placed in a centrifuge for 2 minutes, and the supernatant can be directly used as a qPCR reaction template.

[0064] Further, in step S3, the centrifugal rotation speed of the centrifuge is 10000 r / min. In step S3, the amount of template added is 20% ...

Embodiment 3

[0066] This embodiment provides a detection method of blood direct expansion probe method qPCR:

[0067] Further, the probe method qPCR mix used 2×TaqManFast qPCR Master Mix of Shanghai Sangong No. B639274.

[0068] S4, configure the probe method qPCR reaction system as shown in Table 1 below:

[0069] Table 1 qPCR reaction system.

[0070]

[0071]

[0072] S5, the probe method qPCR reaction program is shown in Table 2:

[0073] Table 2 qPCR reaction program

[0074]

[0075] Further, the ROX used mainly includes two concentrations, ROX Reference Dye (L) and ROX Reference Dye (H), which are selected according to different models of fluorescence quantitative PCR instruments.

[0076] Further, in the described step S4, ROX Reference Dye needs to be selected according to the model of the instrument: ROX Reference Dye (H) suitable model is ABI Prism7000 / 7300 / 7700 / 7900, Eppendorf Realplex4, ABI Step One, ABI Step One Plus; ROX Reference Dye(L) suitable models are ABI ...

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Abstract

The invention discloses a nucleic acid release liquid, a kit and a nucleic acid release method, and relates to the technical field of biology. The nucleic acid release liquid comprises Tris-HCl, ammonium chloride, KOH, Tween-20 and water, the concentration of each component is as follows: Tris-HCl with the concentration of 40-60 mM, ammonium chloride with the concentration of 100-200 mM and KOH with the concentration of 30-50 mM, and the Tween-20 accounts for 0.02%-0.2% of the total volume of the nucleic acid release liquid. When the nucleic acid release liquid is used for treating a sample, genome DNA with stable quality can be obtained without specific protein removal, RNA removal and secondary metabolite removal steps or organic solvent extraction, and the nucleic acid release liquid can be directly used in the fields of PCR amplification, detection, identification and the like without subsequent purification steps. The nucleic acid release liquid has the technical advantage of short time consumption in the operation process, and is more convenient for scientific research personnel or medical personnel in a laboratory to use.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular, to a nucleic acid release solution, a kit and a method for nucleic acid release. Background technique [0002] Whole genome DNA (genomic DNA, gDNA) is the entire genetic information including coding and non-coding sequences in haploid cells. All life information can be found from whole genome DNA, including gene expression, gene mutation and single nucleotide polymorphism closely related to disease. Blood samples have unusual reference value for clinical diagnosis and research. Hematology, genetics and biochemistry all require blood samples to identify and judge various human diseases. Therefore, the isolation of genomic DNA from blood is the basis for clinical diagnosis and genomic research. [0003] There are many PCR reaction inhibitors in whole blood, such as heme, proteins, salts, anticoagulants, etc., making it impossible to directly use whole blood for qPCR / RT-qPCR rea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806C12N15/10C12Q1/686
CPCC12Q1/6806C12N15/1003C12Q1/686C12Q2527/125C12Q2523/32C12Q2563/107
Inventor 赵斯斯薛茜
Owner 生工生物工程(上海)股份有限公司
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