Qiagen plasmid purification handbook 082003 9 product use limitations qiagen, qiafilter, and endofree plasmid kits are developed, designed and sold for. 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. 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. The minelute reaction cleanup kit provides spin columns, buffers, and collection tubes for silica membranebased. With the qiaex ii gel extraction kit, extraction and purification of dna fragments are based on solubilization of agarose and selective, quantitative adsorption of nucleic acids to the qiaex ii silicagel particles in the presence of high salt. Pcr purification kit qiagen protocol qiaquick pcr purification kit. A pancoronavirus rtpcr assay for detection of all known coronaviruses. 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. I am currently using the qiaquick gel extraction kit by qiagen, which is not very efficient and i have to load a lot of pcr product. Can we use the qiagen pcr purification kit after dissolving gel piece for extraction of dna from agarose gels. Elution of the dna is accomplished with a lowsalt solution such as tris buffer or water. Dna fragments from 4 kb10 kb should be purified using the qiaquick gel extraction kit and dna fragments smaller than 70 bp or larger than 10 kb should be extracted with the qiaex ii gel extraction system specifications elution with just 10 l tris buffer or water highly concentrated purified dna dna fragments extracted in 18 minutes.
A detailed protocol for using qiaquick spin columns on the qiacube is provided with the qiacube. Miniprep, qiaquick pcr cleanup kit, minelute gel extraction kits qiagen, 27104, 28104. 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. Poor yield with gel extraction kit for about the umpteenth time now, i have gotten poor yield isolating my cut fragment from a 1% agarose gel. Removal of dye terminators in sanger sequencing reactions. Banerjee lab ucla molecular, cell and developmental biology. 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.
Structure prediction, functional characterization of. Qiaquick gel extraction kit the qiaquick gel extraction kit cat. Then perform gel extraction cleanup with the qiaquick gel extraction kit. Gel extraction kit and a comparable kit from company q following manufacturers recommended protocols. Recoveries of dna fragments in the size range between removed and recovered are not defined. Qiaquick gel extraction kit 250 for gel extraction or cleanup of 250 reactions. The protocol presented here includes steps from constructing a new recombinant hiv1 clone to performing qiaprep spin miniprep kit, qiagen, 27104. Gel extraction with qiaquick gel extraction kit qiagen library preparation with nextera xt dna library prep kit illumina. For more information, please refer to the qiaquick spin handbook, march 2008.
Qiagen plasmid purification handbook 082003 3 contents kit contents 5 storage 8 technical assistance 8 product warranty and satisfaction guarantee 8 product use limitations 9 safety information 9 introduction 10 comparison of qiagen largescale plasmid purification options 10 principle and procedure. The innovative qiacube uses advanced technology to process qiagen spin columns, enabling seamless integration of automated, lowthroughput sample prep into your laboratory workflow. The minelute pcr purification kit and minelute gel extraction kit can be fully automated on the qiacube. Jul 22, 2009 currently im using the qiagen qiaquick gel extraction kit and taking readings after purification with that. Qiagen plasmid purification handbook harvard university. We use qiagens qiaquick gel extraction kit for this purpose. This offer is valid until december 31, 2019 and only when order is placed online through qiagen web shop, your organizations purchasing system, your sales rep, or by email using the cyber19 promo code. For sequencing, add 5 l of purified amplification product to 4 lofabiprism bigdye ready reaction mix and 1 lofa5 m solution of forward or reverse primer. Purified dna was cloned into the plasmid vector, ptz57rt using instaclone pcr cloning kit appendix 1, thermo scientific by few modifications. 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. Agarose gel analysis of the purification procedure 41 appendix b. Basically i was just wondering what types of readings other people get after gel extractions, and if this could have a huge impact on downstream applications such as the ligations im doing.
For cleanup of other enzymatic reactions, follow the protocol as described for pcr samples or use the minelute reaction. The kit is suitable to extract and purify dna of 70 bp to 10 kb from. Electrophoretic analysis of recovered dna from an agarose gel compared to the control. Mar 30, 2007 extraction of pure dna from agarose is essential for many downstream assays including cloning and radioisotopic or fluorescent dna sequencing. The minelute gel extraction kit removes nucleotides, enzymes, salts, agarose, ethidium bromide, and other impurities from dna samples, delivering highly concentrated dna suitable for a variety of downstream applications see figure higher dna concentrations.
If detecting fluorescently labeled multiplex pcr products under denaturing conditions e. Qiaprep miniprep kits use silicagelmembrane technology to eliminate the cumbersome steps associated with loose resins or slurries. Technical information the competent cells provided with the cold fusion cloning kit has the following genotype. Can we use the qiagen pcr purification kit after dissolving.
In this stepbystep dneasy visual protocol, youll see how to extract genomic dna from various samples in just 20 minutes. Qiaex ii handbook qiaex ii agarose gel extraction protocol. Qiagen qiaamp dna micro kit, 50 preps, 1 to 100l sample, 20 to 100l elution volume, whole blood sample, silica technology, manual processing, spin column format, genomic dna, mitochondrial dna purification, 30 min. For more information, please refer to the qiaquick spin handbook, which can be. Automated dna cleanup the qiaquick pcr purification kit and qiaquick gel extraction kit can be fully automated on the qiacube. Qiaquick gel extraction kit protocol 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. Does somebody know a better gel extraction kitprotocol. For purification of genomic and mitochondrial dna from small samples. Qiaquick gel extraction kit protocol using a microcentrifuge.
Qiagen plasmid purification handbook 112005 3 contents kit contents 4 storage 5 quality control 5 product use limitations 6 product warranty and satisfaction guarantee 6 technical assistance 6 safety information 7 introduction 8 principle and procedure 8 equipment and reagents to be supplied by user 10 important notes 11 protocols plasmid or. The minelute qiaquick gel extraction kit provides spin columns for gel extraction. Why is the concentration of dna very low after a gel extraction. This protocol is designed to purify single or doublestranded dna fragments from pcr and other enzymatic reactions see page 8. Qiaquick gel extraction kit protocol syracuse university. Agarose gel analysis of the purification procedure 60. 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 10micro g.
Step 2 running the preparative agarose gel to separate the prepared vector and insert bands from unwanted material, such as incompletely digested dna, enzymes, and other small molecules, run a preparative agarose gel. Qiaquick spin handbook for qiaquick pcr purification kit. The following risk and safety phrases apply to components of. Qiaquick gel extraction kit 50, 28704 from qiagen labsave. Qiaquick gel extraction kit from qiagen biocompare product.
Qiaquick spin handbook qiaquick pcr purification kit for purification of pcr products, 100 bp to 10 kb qiaquick nucleotide removal kit for oligonucleotide 1740mers and dna 40 bp to 10 kb cleanup from enzymatic reactions qiaquick gel extraction kit for gel extraction or cleanup of dna 70 bp to 10 kb from enzymatic reactions. For gel extractioncleanup of up to 10 micro g dna 70 bp to 10 kb from enzymatic reactions. Minelute gel extraction kit for gel extraction of dna fragments 70 bp to 4 kb. The innovative qiacube uses advanced technology to process qiagen spin columns, enabling seamless integration of automated. Qiagen qiaquick gel extraction kit 28704 and 28706. It has not been thoroughly tested and optimized by qiagen. Add 5l of loading dye to your pcr and load the entire volume onto the gel. 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. Qiagen research kits compatible with the qiavac 24 plus application kit accessories plasmid minipreps qiaprep spin miniprep kits dna cleanup qiaquick pcr purification kits qiaquick gel extraction kits qiaquick nucleotide removal kits minelute pcr purification kits.
Using the small gel rig, pour a 1% low melting point agarose gel 0. A260230 readings post gel extraction general lab techniques. Its probably not too important during most of the procedure, but its worth bearing in mind at the elution step. Qiaprep miniprep handbook washington state university. Minelute gel extraction kit 50 from qiagen selectscience. Construc tion and charac terization of est libraries from. Background using silver staining purify pcr products before loading onto the gel e. Qiavac vacuum manifolds 30 handling guidelines for qiavac 24 31 qiavac 6 32 ordering information 33 qiagen companies and distributors 35 qiaquick spin handbook 032001 3. Qiaquick spin handbook 032008 19 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. Dna ladder was run on a 2% agarose gel and 4 different fragment sizes 200 bp, 500 bp, 1 kb, and 5 kb were recovered using omega bioteks e. 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. A pancoronavirus rtpcr assay for detection of all known.
Up to 400 mg agarose can be processed per spin column. For departmental related matters, please visit the department of molecular, cell, and developmental biology website. Handbook 1 1 qiaquick gel extraction kits 50 250 catalog no. Apr 08, 2008 if you use qiagens gel extraction kit, you may have noticed that after each spin, a small droplet tends to form inside the column at the side that was to the outside of the centrifuge rotor due to the shape of the column. For more information, please refer to the qiaquick spin handbook. Using a microcentrifuge or vacuum manifold, dna ranging from 70 bp to 10 kb is purified from 124 samples. Handbook qiagen plasmid mini, midi, maxi, mega, and giga kits. Dna ranging from 70 bp to 10 kb is purified using a simple and fast bindwashelute procedure and an elution volume of 3050. Qiaquick gel extraction kit 250 from qiagen selectscience. I put in 1ug and i am getting back about 10nul but, my 260280s are jumping all over the place, as is my concentrations. Qiagen plasmid purification handbook 082003 5 kit contents. Please be sure to read the qiaquick spin handbook and the detailed qiaquick gel.
852 164 1069 1185 761 775 1510 326 687 84 6 1119 113 956 1551 981 1015 435 1281 105 1177 591 83 260 1404 860 794 1360 1446 1640 221 49 1386 1307 1466 826 1360 1349 1435 1247 1126 223