Omega胶回收试剂盒操作

Benefits
The E.Z.N.A.® Gel Extraction Kit means:
Related Products Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
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Kit Contents
Product Number D2500-00 D2501-00 D2502-00
5 5 5 5 ml 1 ml 5 ml 1
D2500-01 D2501-01 D2502-01
50 50 50 40 ml 10 ml 25 ml 1
D2500-02 D2501-02 D2502-02
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Benefits . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Binding Capacity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 Kit Contents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Materials Supplied By User . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Gel Extraction Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Optional Vacuum Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Trouble Shooting Guide . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 Short Protocol For Experienced Users . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
百度文库Contents
Introduction
The E.Z.N.A.® family of products is an innovative system that radically simplifies extraction and purification of nucleic acids from a variety of sources. Key to the system is the new HiBind® matrix that specifically, but reversibly, binds DNA or RNA under certain optimal conditions, allowing proteins and other contaminants to be removed. Nucleic acids are easily eluted with Elution Buffer provided. Gel purification of DNA is a common technique for isolation of specific fragm ents from reaction mixtures. However, m ost methods either fail to com pletely rem ove agarose (which can lead to problem s in downstream manipulations), shear the DNA, or result in very low yields. The E.Z.N.A.® Gel Extraction Kit uses HiBind® technology to recover DNA bands 50 bp-40 kb from all grades of agarose gel in yields exceeding 85%. The DNA band of interest is excised from the gel, dissolved in Binding Buffer, and applied to a HiBind ® DNA spin-column. Following a rapid wash step, DNA is eluted with Elution Buffer and is ready for other applications. The product is suitable for ligations, PCR, sequencing, restriction digestion, or various labeling reactions.
Revised March 2005
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Speed - DNA recovery from agarose in <15 m in Reliability - Optimized buffers guarantee pure DNA Safety - No organic extractions Quality - Purified DNA is suitable for any application
New in this edition
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pH indicator has been introduced. New DNA Elution Buffer is supplied. New SP W Buffer has been introduced
Binding Capacity
Each HiBind ® DNA column can bind ~25 :g DNA.
200 200 200 1 50 m l 20 ml 3 x 25 ml 1
Gel Extraction Protocol
Please read this booklet thoroughly to ensure that you are familiar with the entire procedure. E.Z.N.A.® Kits are designed to be simple, fast, and reliable, provided that all steps are followed diligently. Up to 400mg agarose gel can be processed per spin column. 1. Perform agarose gel electrophoresis to fractionate DNA fragm ents. Any type or grade of agarose may be used. It is strongly recommended, however, that fresh TAE buffer or TBE buffer be used as running buffer. Do not re-use running buffer as its pH will increase and reduce yields. W hen adequate separation of bands has occurred, carefully excise the DNA fragment of interest using a wide, clean scalpel. Determine the approximate volume of the gel slice by weighing it in a clean 1.5 ml microfuge tube. Assuming a density of 1 g/ml of gel, the volume of gel is derived as follows: A gel slice of m ass 0.3 g will have a volume of 0.3 ml. Add equal volume of Binding Buffer (XP2). Incubate the mixture at 55o C60o C for 7 min or until the gel has completely melted. Mix by shaking or inverting the tube every 2-3 minutes. Centrifuge the tube briefly to collect all the liquid to the bottom of the tube. Note: For DNA fragment less than 500bp, add 1 sample volume of isopropanol after the addition of Binding Buffer (XP2). Important: Monitor the pH of the Gel/Binding Buffer mixture after the gel completely dissolves. DNA yield will significantly decrease when pH > 8.0. If the color of the mixture become orange or red, Add 5 :l of 5M sodium acetate, pH 5.2, to bring the pH down. After this adjustment, the color of the gel/Binding Buffer mixture should be light yellow. 4. Apply up to 700 :l of the DNA/agarose solution to a HiBind ® DNA spincolumn assem bled in a clean 2 ml collection tube (provided) and centrifuge in a microcentrifuge at 8,000-10,00 0 x g for 1 min at room temperature. Discard the liquid. Re-use the collection tube in Steps 5-8. For volumes greater than 700 :l, load the colum n and centrifuge successively, 700 :l at a time. Each HiBind ® spin-column has a total capacity of 25-30 :g DNA. Discard liquid and add 300:l Binding Buffer. Centrifuge at 10,000 x g for 1 minutes. Add 700 :l of SPW Buffer diluted with absolute ethanol into the column and wait 2-3 minutes. Centrifuge at 10,000 x g for 1 min at room temperature to wash the sample. Optional: Discard liquid and repeat step 6 with another 700 :l SPW Buffer. Note: Perform this second wash step for any salt sensitive downstream applications. 8.