Dna extraction from agarose gels matt lewis, department of pathology, university of liverpool very nice protocol which covers three methods of extracting dna from agarose gel. Thermo scientific genejet gel extraction kit fisher scientific. For more information, please refer to the qiaquick spin handbook. The qiaex ii gel purification kit can be used for extraction and purification of dna from either tae or tbe agarose or polyacrylamide gels. Jul 20, 2009 the qiaex ii gel purification kit can be used for extraction and purification of dna from either tae or tbe agarose or polyacrylamide gels. The qiaquick gel extraction kit enables removal of nucleotides, enzymes, salts, agarose, ethidium bromide, and other impurities from samples, ensuring up to 80% recovery of dna see figure high recoveries from gels. The input amount of dna to be purified should not exceed the binding capacity of the column 5. The innovative qiacube uses advanced technology to process qiagen spin columns, enabling seamless integration of automated, lowthroughput sample prep into your laboratory workflow. Qiaamp mini and qiaamp minelute spin columns are processed on the. This protocol is designed to extract and purify dna of 70 bp to 10 kb from standard or lowmelt agarose gels in tae or tbe buffer. Gel extraction protocol qiaquick gel extraction kit.
Marker qiagen kit brain liver heart purelink kit qiagen kit purelink kit qiagen kit purelink kit marker figure 5. The minelute pcr purification kit and minelute gel extraction kit can be fully automated on the qiacube. Qiagentips should be stored dry and at room temperature 1525c. Qiaquick pcr purification kit using a microcentrifuge 22. Briefly place the excised gel slice on absorbent toweling to remove residual buffer. This can be achieved by using a wider gel comb and running the gel at a. For departmental related matters, please visit the department of molecular, cell, and developmental biology website. However, with 500 ng we ultimately get 100 ng for a 20% yield.
Gel extraction protocol qiaquick gel extraction kit protocol. Qiaquick gel extraction kit 50 from qiagen sample to insight. The kit was tested in the extraction of dna fragments from an agarose gel according to the protocol described in the manual. For cleanup of other enzymatic reactions, follow the protocol as described for pcr samples or use the minelute reaction cleanup kit. Isolate a suitable piece of tissue and place in a uvcrosslinked 1.
For storage longer than 12 months or if ambient temperatures constantly exceed 25c, qiagen protease should be stored dry at 28c. Protocol qiaquick spin handbook 032001 23 qiaquick gel extraction kit protocol using a microcentrifuge this protocol is designed to extract and purify dna of 70 bp to 10 kb from standard or lowmelt agarose gels in tae or tbe buffer. Qiaex ii handbook qiaex ii agarose gel extraction protocol. After adding rnase a, buffer p1 should be stored at 28c and is stable for 6. Laboratory protocol for dna extraction from oracollectdx ocd. Protocol for extraction and purification of genomic dna from. Reconstituted qiagen protease is stable for 2 months when stored at 28c. For maximum convenience and value, columns and buffers are also available separately.
These are available online in convenient and compact pdf format. Each fragment was manually excised from the agarose gel and processed. Laboratory protocol for dna extraction from oracollectdx. The qiaquick gel extraction kit provides spin columns, buffers, and collection tubes for silicamembranebased purification of dna fragments from gels up to 400 mg slices or enzymatic reactions. Qiaquick gel extraction kit using a vacuum manifold. Quickstart protocol 11 rneasyprotectminikit 50 250 catalogno. They can be stored for at least 2 years without showing any reduction in performance, capacity, or quality of separation. All restriction enzymes, taq dna polymerase, 1kb dna ladder, and t4 dna ligase are obtained from promega madison, wi. Protocol qiaquick spin handbook 032001 23 qiaquick gel extraction kit protocol using a microcentrifuge this protocol is designed to extract and purify dna of 70 bp qiaquick gel extraction kit for gel the same steps as the manual qg and leads to inefficient dna binding. Banerjee lab ucla molecular, cell and developmental biology. Qiagen plasmid kits should be stored at room temperature 1525c. For gel extractioncleanup of up to 10 g dna 70 bp to 10 kb from enzymatic reactions. Prestotm 96 well gel extraction kit protocol please read the entire instruction manual prior to starting the protocol procedure. The maximum amount of gel per spin column is 400 mg.
Thebolded should benoticed foranice dna extraction. Qiaex ii gel extraction kit from qiagen biocompare product. Qiagen qiaquick gel extraction kit 28704 and 28706. Recoveries of dna fragments in the size range between removed and recovered are not defined. Cut asclose tothe dna aspossible tominimize thegelvolume. Minelute handbook 062004 11 adsorption to minelute membrane salt and ph dependence the minelute silicagel membrane is uniquely adapted to isolate dna from both aqueous solutions and agarose gels, and up to 5 g dna can bind to each minelute column.
Protocol qiaquick spin handbook 072002 23 qiaquick gel extraction kit protocol using a microcentrifuge this protocol is designed to extract and purify dna of 70 bp to 10 kb from standard or lowmelt agarose gels in tae or tbe buffer. Qiaquick pcr purification kit protocol using a microcentrifuge this protocol is designed to purify single or doublestranded dna fragments from pcr and other enzymatic reactions see page 8. Qiaex ii gel extraction kit from qiagen biocompare. Lyophilized qiagen protease can be stored at room temperature for up to 12 months without any decrease in performance. Using a qiagen gel extraction kit, you will dissolve each gel piece, resulting. Qiaquick nucleotide removal kit protocol 21 qiaquick gel extraction kit protocols 23 using a microcentrifuge 23 using a vacuum manifold 25 troubleshooting guide 28 appendix. I was curious what else has been tried for gel extraction. We use the standard protocol from qiagen, gel extraction, dissolve in qg buffer at 42c and purify via anion exchange columns. Excise the agarose gel slice containing the dna fragment of interest with a clean, sharp scalpel under ultraviolet illumination. May 24, 2010 gel extraction is a technique used to isolate a desired fragment of intact dna from an agarose gel following agarose gel electrophoresis. Excise gel slice containing thedna fragment using aclean scalpel orrazor blade. Improving gel extraction yields biology stack exchange. Add ethanol 96100% to buffer pe before use see bottle label for volume. Gel extraction is a technique used to isolate a desired fragment of intact dna from an agarose gel following agarose gel electrophoresis.
This protocol is designed for purification of total dna from grampositive bacteria. This protocol is designed to extract and purify dna of 70 bp to 10 kb from standard or low melt agarose gels in tae or tbe. Dna fragments are excised from an agarose gel and are diluted by addition of four volumes of gel dissolving buffer. For departmental related matters, please visit the department of molecular, cell, and developmental biology website phone. For departmental related matters, please visit the department of. Follow the agarose gel electrophoresis protocol with the following amendments note.
Dna ranging from 70 bp to 10 kb is purified using a simple and fast bindwashelute procedure and an elution volume of 3050. Place each gel piece into an empty microcentrifuge tube, throw away the gel into the gel waste and the razor into the sharps container, clean the gel doc surface, and return to the lab with your tubes containing the gel fragments, the lab coat, and gel photo. These are available online in convenient and compact pdf format at. L of buffer al to the cell suspensionproteinase k and vortex for 15 seconds. Follow the agarose gel electrophoresis protocol with the following amendments. Protocol for extraction and purification of genomic dna.
Lyophilized qiagen protease can be stored at room temperature 1525c for up to. A mixture of 7 dna fragments ranging from 10 kb down to 0. Qiaquick gel extraction kit protocol using a microcentrifuge. Up to 400 mg agarose can be processed per spin column. Qiagen plasmid purification handbook harvard university. The binding buffers in minelute spin kits provide the correct salt concentration. Using a microcentrifuge or vacuum manifold, dna ranging from 70 bp to 10 kb is purified from 124 samples. Monarch dna gel extraction kit reproducibly recovers dna over a broad range of molecular weights. Monarch nucleic acid purification kits are optimized for maximum performance and minimal environmental impact.
For more information, please refer to the qiaquick spin handbook, march 2008. Protocol qiaex ii agarose gel extraction protocol this protocol is designed for the extraction of 40bp to 50kb dna fragments from 0. Incubate at 50c for 10 min or until the gel slice has completely dissolved. Visual inspection of rna integrity by agarose gel electrophoresis. The following protocol for sds removal was developed for cytoplasmic intermediate filaments, and we successfully applied it to lamin a. This can be achieved by using a wider gel comb and running the gel at a lower voltage. Product information thermo scientific genejet gel extraction kit.
Cut as close to the dna as possible to minimize the gel volume. Dna extraction from the gel using centrifuge step procedure 1 excise gel slice containing the dna fragment using a clean scalpel or razor blade. Dna adsorbs to the dneasy membrane in the presence of high concentrations of chaotropic salt, which remove. Qiaamp dna mini kit and qiaamp dna blood mini kit handbook. The kit utilizes a proprietary silicabased membrane technology in the form of a convenient spin column. Youll be thrilled to pieces do you need a faster, more reliable solution for dna fragmentation and library construction. Add 3 volumes buffer qg to 1 volume gel 100 mg gel 100. Be sure and close the bottle tightly after each use to. Transfer the gel slice to a piece or plastic wrap or a weighing boat.
Gel purification is most efficient with lower % agarose gels, so you will want to stay in the 0. If you have questions about any of the information in the bench guide, or about molecular biology in general, please call your local qiagen technical service department or your local distributor see back cover. Qiaquick gel extraction kit 50 from qiagen sample to. Processing nondsp qiaamp mini and qiaamp minelute spin columns on the qiavac 24 plus a vacuum pump capable of producing a vacuum of at least 900 mbar e. Thermo scientific genejet gel extraction kit is designed for rapid and efficient purification of dna fragments from standard or lowmelting point agarose gels run in either tae or tbe buffer. The extraction process is designed such that phenol or ethanol precipitates are not required, instead making use of silica particles to enhance recovery of very small or large dna fragments. Qiavac vacuum manifolds 30 handling guidelines for qiavac 6 31 qiavac 24 32 ordering information 33 qiagen companies and distributors 35 qiaquick spin handbook 072002 3. The polymerase chain reaction pcr purification kit, qiaex ii gel extraction kit, qiaamp tissue kit, and dna miniprep spin kit are from qiagen santa clarita, ca. What is the best gel extraction method for a high dna. A classic method is to crush the gel mortar and pestle, elute in triethanolamine bicarbonate ph7, apply to a c18 column, wash with buffer, wash with buffer in 10% acetonitrile, and elute in tea. Qiaquick pcr purification kit using a microcentrifuge. Different net proteins and fragments will bind sds. This protocol is designed to purify single or doublestranded dna fragments from pcr and other enzymatic reactions see page 8.
Add absolute ethanol see the bottle label for volume to wash buffer then mix by shaking for a few seconds. Forceps are to be sterilized in fine science tools heat block at. Sigma gel extraction kit is best for dna extraction and purification from gels. The protocol describes the preliminary harvesting of bacteria and incubation with lysozyme to lyse their cell walls before dna purification. Qiaquick gel extraction kit for extraction of dna fragments 70 bp 10 kb from. Paper strip method, spincolumns and dialysis tubing semipermeable membrane, visking tubing. Figure 4, evaluated for integrity by both gel electrophoresis figure 5, and analyzed on the 2100 bioanalyzer instrument using the rna 6000 nano kit figure 6. Qiagen qiaquick gel extraction kit, 50 rxns, 30 to 50l elution volume, 10g binding capacity, dna sample, tube format, silica technology, manual processing, 70 bp to 10 kb fragment, fast and convenient procedure, for gel extractioncleanup of up to 10g dna 70.
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