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DNA gel extraction reagent and method for purifying PCR products by using reagent

A technology for products and glue blocks, applied in the field of biological reagents, can solve problems such as environmental pollution and waste, and achieve the effects of protecting the environment, reducing garbage, saving social resources and use costs

Inactive Publication Date: 2018-11-13
WUHAN UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The technical problem to be solved by the present invention is: how to provide a DNA gel extraction reagent to solve the problems of environmental pollution and waste caused by the one-time use of the existing DNA gel extraction kits

Method used

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  • DNA gel extraction reagent and method for purifying PCR products by using reagent

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1DNA gel extraction uses the configuration of agarose gel solution

[0025] The agarose gel solution includes 6 mol / L Guanidine.HCl, 25 mmol / L Sodium Citrate, 0.5% N-Lauroylsarcosine, and 0.1 mol / L 2-mercaptoethanol.

[0026] Taking 100 ml of agarose gel solution as an example, the preparation method is as follows:

[0027] (1) Weigh 57.3 grams of guanidine hydrochloride, 7.35 grams of sodium citrate and 0.5 grams of lauroyl sarcosine respectively in an electronic balance, place the medicine in a beaker with a capacity of 250 milliliters, add 50 milliliters of distilled water, and place on a magnetic stirrer Heat to dissolve.

[0028] (2) Take 6.8 microliters of 2-mercaptoethanol with a 20 microliter pipette gun, add it to the above solution, stir and mix well.

[0029] (3) Transfer the solution to a 100ml graduated cylinder, add distilled water to make it 100ml. Divide the volume-contained solution into 50 ml sterile centrifuge tubes, and seal them for st...

Embodiment 2

[0030] Embodiment 2 configuration of washing liquid

[0031] The washing liquid contains 10mmol / L Tris.HCl (pH 8.0), 100mmol / L NaCl, 1mmol / L EDTA, 80% ethanol.

[0032] Take 100ml washing liquid as an example below, the preparation method is as follows:

[0033] (1) Prepare 1mol / L Tris.HCl (pH 8.0) mother liquor and 0.5mol / L EDTA mother liquor according to conventional methods;

[0034] (2) Weigh 0.585 gram of NaCl in electronic balance, place in 250 milliliter reagent bottle;

[0035] (3) Add 1 ml of 1mol / L Tris.HCl (pH8.0) mother solution and 200 microliters of 0.5mol / LEDTA mother solution to the reagent bottle;

[0036] (4) Add 18.5 milliliters of distilled water to completely dissolve NaCl in the reagent bottle;

[0037] (5) Measure 80 ml of absolute ethanol with a graduated cylinder, add it to the reagent bottle, mix thoroughly with the solution, and seal it with a cap.

experiment example

[0039] In order to identify the effect of the DNA gel extraction reagent of the present invention on DNA gel extraction or purification, utilize the regenerated silicon column in the Qiagen and Axygen gel extraction kits (the preparation method of the regenerated silicon column refers to the patent application number: 201710437190.7, and the name of the invention is : silicon column rapid regeneration method) and reagent of the present invention are purified to the DNA gel after electrophoresis; Simultaneously compare with the DNA purification effect of Qiagen and Axygen DNA gel extraction kit original silicon column and reagent.

[0040] Utilize reagent of the present invention to complete DNA gel extraction method as follows:

[0041] (1) Prepare 1% agarose gel according to the usual method. After the gel is cooled, pull out the loading comb and place it in the electrophoresis tank containing 1xTAE, load 50 microliters of PCR product (LIF DNA) in the loading hole, Perform el...

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Abstract

The invention discloses a DNA gel extraction reagent and a method for purifying PCR products by using the reagent, wherein the DNA gel extraction reagent includes an agarose gel dissolving solution for extracting DNA gel and a scrubbing solution. A formula of the agarose gel dissolving solution for extracting the DNA gel comprises 6 mol / L of guanidine hydrochloride, 25 mmol / L of sodium citrate, 0.5% of lauroyl sarcosine, and 0.1 mol / L of 2-mercaptoethanol; a formula of the scrubbing solution comprises 10 mmol / L of Tris.HCl, 100 mmol / L of NaCl, 1 mmol / L of EDTA, and ethanol with the volume concentration of 80%, wherein the pH value of Tris.HCl is 8.0. The DNA gel extraction reagent can be used together with a regenerated silicon column, can reduce the use amount of disposable commercial kits, reduces the generation of laboratory waste, protects the environment, and saves social resources and use costs.

Description

technical field [0001] The invention belongs to the field of biological reagents, in particular to a DNA gel extraction reagent and a method for purifying PCR products using the reagent. Background technique [0002] Gene cloning (also known as molecular cloning) is the most commonly used technique in modern molecular biology, widely used in biology, medicine, biopharmaceutical industry and other fields. Gene cloning refers to the process of recombining the obtained target gene or DNA (deoxyribonucleic acid) fragment with the plasmid (vector) in vitro and introducing the recombinant vector into the host for replication, and obtaining a large amount of recombinant DNA by extracting the plasmid. This process includes amplification and purification of the target gene, restriction enzyme digestion reaction, agarose electrophoresis and DNA gel extraction or purification of the digested product, ligation of the target DNA with the plasmid, bacterial transformation of the ligated p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1006
Inventor 邓文生张程赵沙沙
Owner WUHAN UNIV OF SCI & TECH
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