Thermostable β-Agarase
- Manufacturer :
- Nippon Gene Co., Ltd.
- Storage Condition :
- Keep at 2-10 degrees C.
- Structural Formula
- Label
- Packing
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Comparison
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Certificate of Analysis
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30U
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In stock in Japan |
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300U
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In stock in Japan |
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※Check availability in the US with the distributor.
Document
- Package Insert
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- Spectral Data
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- Certificate of Analysis
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- Calibration Certificate
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- Analytical Charts
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Simple Protocol
Data
Cloning of the DNA recovered from agarose gel pieces
The 500bp DNA fragment derived from λDNA was amplified by PCR.
PCR product were separated by 3% Agarose 21 (low molecular weight separation; NIPPON GENE).
A required band was excised from the gel with a razor.
After melting gel piece, it was treated by Thermostable β-Agarase.
9 µl of the gel solution was added to 1 µl of cloning vector DNA (3 kbp, 50 ng/µl) to set up ligation reaction using Ligation-Convenience Kit (NIPPON GENE).
After ligation reaction, competent cell were transformed and then the number of colonies were counted. As the control, DNA solution purified by ethanol precipitation was used for ligation reaction.
Cloning of the DNA recovered from agarose gel pieces
T4 GT7 DNA (166 kbp), λDNA (48.5 kbp), pUC19 DNA (2.69 kbp) separated by agarose gel electrophoresis in 1×TAE was excised from 0.3% Agarose L (low-melting point; NIPPON GENE). The gel pieces were treated by Thermostable β-Agarase or spin column and the collected amount of DNA was compared. One third amount of DNA was separated using 0.3% Agarose H (high gel strength; NIPPON GENE).
How to use
How amount of Thermostable β-Agarase should be used?
Generally, add the enzyme at a ratio of 6 unit (6µl) per 200 mg agarose gel. The amount of the enzyme can sometimes be reduced by the density of the gel and the condition.
- ▼The amount of Thermostable β-Agarase which is needed for agarose gel to be hydrolyzed in 5 minutes.
Agarose conc. Enzyme volume 1.0% Agarose S 2.0 µl (2.0 unit) 1.0% Agarose S 3.0 µl (3.0 unit) 2.0% Agarose S 5.0 µl (5.0 unit) 1.5% Agarose XP 3.0 µl (3.0 unit) - ▼The amount of Thermostable β-Agarase which is needed for agarose gel to be hydrolyzed in 10 minutes.
Agarose conc. Enzyme volume 1.5% Agarose S 1.5 µl (1.5 unit) 2.0% Agarose S 3.0 µl (3.0 unit) 3.0% Agarose 21 5.5 µl (5.5 unit)
FAQ
About the product
- How can it be confirmed whether gel is completely hydrolyzed?
- When not gelatin even if DNA solution is cooled on the ice, agarose gel is completely hydrolyzed.
- What is an advantage of thermostability?
- It is not only used for standard gel, but can be used for low-melting pint gel. Reaction finishes in only 10 minutes.
- How much is recovery efficiency of a DNA?
- Almost all of DNA can be recovered in general.
- Thermostable β-Agarase maintains the enzyme activity at a long term?
- This enzyme is stable for 2 years at 4 – 20℃. When keeping in the room temperature, it is stable for 1 year. The activity is not altered during one-hundred cycles of freezing and thawing.
- What amount of hydrolyzation solution produces?
- When using 200 mg Agarose gel, the amount of hydrolyzed solution will be 200 µl.
- How should hydrolyzed solution be concentrated?
- DNA can be concentrated be ethanol precipitation.
- Can hydrolyzed solution be used for in vitro transcription reaction?
- It is possible to synthesize RNA by using hydrolyzed solution directly as the template. CUGA®7 in vitro transcription Kit (NIPPON GENE) was used for RNA synthesis.
- Can hydrolyzed solution be used for DNA sequencing?
- It is possible to sequence by using hydrolyzed solution directly as the template.
- Is it possible to recover RNA from denaturing agarose gel by using Thermostable β-Agarase?
- It can be used, however formaldehyde inhibits the enzyme activity. Therefore, it’s necessary to increase the amount of the enzyme.
Overview / Applications
| Outline | This product is for research use only. Do not administer it to human. for Recovery of nucleic acids from agarose gels β-agarase is an enzyme which hydrolyzes β-1,4 linkages in agarose to produce neoagaro- oligosaccharides. Agarose digested by β-agarase does not gelate again, therefore, nucleic acids can be recovered from agarose gels. This enzyme was isolated from deep-sea microorganism by Japan Agency for Marine-Earth Science and Technology (JAMSTEC) using the submergible SHINKAI 6500. Features:
Procedures 1 |
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Property
Manufacturer Information
Alias
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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