Principle and procedure qiagen plasmid purification protocols are based on a modified alkaline lysis procedure, followed by binding of plasmid dna to qiagen anionexchange resin under appropriate lowsalt and ph conditions. Although the development of plasmid isolation kits utilizing silica spin columns. Favorprep tm plasmid dna extraction mini kit user manual. Genelute plasmid dna miniprep kit 70 purifications gen. Dna binding capacity is tested by determining the recovery obtained with 10 g and 20 g of various plasmid dnas. The mirneasy mini kit is designed for purification of total rna, including mirna and other small rna molecules, from cultured cells and various animal and human tissues. Plasmid isolation miniprep adapted from qiagen miniprep handbook p1. This product is not intended for the diagnosis, prevention, or treatment of a disease. For purification of bacs, pacs, p1s, and purification of double. Plasmid dna isolated with zyppy miniprep kits result in the highest transfection efficiencies. This technology is based on binding dna to silicabased membranes in chaotropic salts, washing dna with ethanolcontained wash buffer. Dna mini prep protocol qiagen spin protocol plasmid prep.
Qiaamp dna mini kit and qiaamp dna blood mini kit handbook. The plasmid concentration was as low as 50ngul measured by fluorescence kit from molecular device. Bac prep using quiagen plasmid mini prep kit do not need spin columns 1. Storage of rna later reagent at lower temperatures may cause precipitation.
Qiagen, qiafilter, hispeed, and endofree plasmid kits, provide optimal. Plasmid dna was purified from bacterial cultures using nucleobond xtra midi, nucleobond xtra maxi, and corresponding midiprep and maxiprep kits from qiagen following the protocols for each kit in each case, the maximum input volume specified by the corresponding kit protocol was processed. Thermo scientific genejet plasmid miniprep kit has been tested in the isolation of puc19 dna from e. Please consult the material safety data sheet msds for information regarding hazards and safe handling. In accordance with qiagen s isocertified total quality management system, each lot of qiaprep miniprep kit is tested against predetermined specifications to ensure consistent product quality. Select a single well isolated colony from your freshly transformed cell culture. Qiaprep spin miniprep kit protocol using a microcentrifuge this protocol is designed for purification of up to 20 g of highcopy plasmid dna from 15ml overnight cultures of liin lb luriabertani medium. Qiagen plasmid mini kit 100, 12125 from qiagen labsave. The kits are designed to efficiently isolate plasmid dna from e.
Centrifugation tubes or vessels with suitable capacity for the volumes specified in the appropriate protocol. For cosmid and lowcopynumber plasmid purification see recommendations on page 12. Maxipreps for plasmid dna purificationnucleobond xtra. Qiagen plasmid purification handbook sierra bermeja. Spin miniprep kit for purification of highquality plasmid dna. We recommend all users of qiagen products to adhere to the nih guidelines that have. Rely on qiagen assay technologies to amplify your samples dna sequence.
General description the genelute plasmid miniprep kit offers a simple, rapid, and costeffective method for isolating plasmid dna from recombinant e. Timerun, 20g yield, 1 samplerun, transfectiongrade plasmid or cosmid dna purification, anionexchange technology, manual processing, ideal for transfection, cloning, sequencing, capillary sequencing, pcr, in vitro. Plasmid dna mini kit i protocol spin protocol all centrifugation should be performed at room temperature unless otherwise noted. Principles behind plasmidprinciples behind plasmid. For purification of lowcopy plasmids and cosmids, large plasmids 10 kb, and dna prepared using other.
The purelink hipure plasmid miniprep kit isolates high yields of pure plasmid dna from e. For qiagen plasmid midi, maxi, mega, and giga kit protocols. Favorprep plasmid extraction mini kit is an excellent tool offering a speed and economic method to purify plasmid dna from bacteria cultures. Precipitate by adding 350 ul of n3 buffer 1020 minutes on ice 6. Blood and body fluid vacuum protocol 30 tissue protocol qiaamp dna mini kit only 33. Qiagen plasmid purification handbook 082003 5 kit contents qiagen plasmid kits qiagen plasmid kit midi 25 midi 100 maxi 10 maxi 25 maxi 100 catalog no. Product use limitations the rneasy mini kit is intended for molecular biology applications. The quality of isolated dna has been evaluated in a set of tests. Quickstart protocol march 2016 qiagen plasmid mini, midi and maxi kits the qiagen plasmid mini kit cat. The isolated plasmid dna is of high purity, equivalent to two. Cultured cells 49 fixed tissues 50 bacteria 52 yeast 54 viral dna 55 eye, nasal, or pharyngeal swabs 56 mitochondrial dna from platelets 57.
Kits, or qiagen products in general, please do not hesitate to. New users and users wanting to purify lowcopy plasmids and cosmids, large plasmids 10 kb, and dna prepared. Purifying plasmid dna from bacterial colonies using the qiagen miniprep kit the jove video player is compatible with html5 and adobe flash. Plasmid dna purification using the qiaprep spin miniprep kit and a. The mirneasy mini kit combines phenolguanidinebased lysis of samples and silica membranebased purification of total rna. Qiagen is the leading provider of innovative sample and assay technologies, enabling. Materials and methods air samples for isolation of fungi presterilized. Product use limitations qiaprep miniprep kits are developed, designed, and sold for research purposes only. Please read important notes before starting following steps. 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. This protocol is designed to isolate plasmid dna from e. Qiagen plasmid plus midi kit 100 from qiagen selectscience. The qiacube is preinstalled with protocols for purification of plasmid dna, genomic dna, rna, viral nucleic acids, and proteins, plus dna and rna cleanup. Qiagen plasmid mini kit 100 from qiagen selectscience.
These are usually called mini, midi and maxi preps. Qiaprep spin miniprep kit using a microcentrifuge this protocol is designed for the purification of up to 20 g highcopy plasmid dna from 15 ml overnight e. Low yield of plasmid extraction by qiagen miniprep. Four different cell lines cos7, hek293, hela, and huh7 were grown to 8090% confluence and transfected with 100 ng of each plasmid, in complex with 0.
These kits come in several sizes that allow the processing of a few g of plasmid all the way up to the mg range. This protocol is designed for purification of up to 20 g of highcopy plasmid dna from. Qiagen plasmid mini kit, 100 preps, 3 to 10ml culture volume, 80 min. The purelink quick plasmid miniprep kit is designed for rapid isolation of sequencinggrade plasmid dna from li in 3045 minutes, using silica membrane columns. The genelute plasmid miniprep kit offers a simple, rapid, and costeffective method for isolating plasmid dna from recombinant e.
The range of protocols available is continually expanding, and additional qiagen protocols. These are available online in convenient and compact pdf format. These are often sold as kits by several different companies and are the most used method of purifying plasmid dna today. Older browsers that do not support html5 and the h.
Qiagen robotic systems are not available in all countries, please inquire. Qiaprep spin miniprep kit highyield protocol english pdf. Highest quality yield, super easy protocol, user friendly instructions and great value for money. Hipure plasmid purification kits allow you to isolate high yields of highly pure plasmid dna. The range of protocols available is continually expanding, and additional qiagen protocols can be. Verify the success of a plasmid dna miniprep using gel electrophoresis. Up to 400 mg agarose can be processed per spin column.
Rneasy mini kit, rneasy protect mini kit, and rneasy plant mini kit is tested against predetermined specifications to ensure consistent product quality. Principles behind plasmidprinciples behind plasmid purifications. The rneasy mini kit, rneasy protect mini kit including rnalater rna. The purelink quick plasmid miniprep kit allows isolation of highquality plasmid dna from 15 ml cultures in 3045 minutes, using the purelink spin columns. The range of protocols available is continually expanding, and additional qiagen protocols can. This kit can also be used for dna cleanup from enzymatic reactions see page 8. Since returning to the lab in 2021 i have tried a purelink hipure plasmid dna purification kit and the qiagen endofree plasmid maxi kit and got less plasmid out of these kits than from the. Timerun, up to 1mg yield, for the fastest, most convenient purification of up to. Luciferase ities were determined in lysates from cells transfected with dna isolated by various methods. Innovative zymopure technology enables elution of ultrapure endotoxinfree plasmid dna in as little as 25 l. Fastreadytouse plasmid dna typically in 3045 minutes flexibleformats work with. Qiagen plasmid purification handbook harvard university. Qiagen plasmid plus midi kit, 100 preps, 25 to 50ml culture volume, transfectiongrade plasmid or cosmid dna purification, advanced silica technology, manual processing, 20 min.
These are available online in convenient and compact pdf format at. The protocol for hispeed midi and maxi kits is optimized for use with cultures grown. Plasmid dna mini kits combine the power of hibind technology with the. Viral rna can be purified from plasma treated with. For purification of up to 10 mg transfectiongrade plasmid or cosmid dna. A rapid and efficient assay for extracting dna from fungi. The important notes, on page 16, and the comments within the qiaamp viral rna mini protocols, beginning on page 27, are particularly valuable. Follow the qiagen protocol if there is confusion or a discrepancy. List the function of each solution used in a plasmid dna miniprep. The rneasy plus mini kit should be stored dry at room temperature 1525. Mix gently by inverting the tube 46 times and incubate at room temperature for exactly 5 min.
Protocols plasmid dna purification using the qiaprep spin miniprep kit and a microcentrifuge 22 and 5 ml collection tubes 23 and a vacuum manifold 24 plasmid dna purification using the qiaprep 8 miniprep kit 28 plasmid dna purification using the qiaprep 8 turbo miniprep kit 31. Qiagen plasmid purificationhandbook 042012 6 data sheets msdss. No fapde 004 fapde 100 fapde 300 favorprep plasmid extraction mini kit fapd1 buffer 1. These products arenotintendedforthediagnosis,prevention,ortreatmentofadisease. Tissue protocol qiaamp dna mini kit only 33 buccal swab spin protocol 37 buccal swab vacuum protocol 39 dried blood spot protocol 41 troubleshooting guide 43 appendix 49 protocols for. Qiaquick gel extraction kit protocol using a microcentrifuge. Compare with other harmful and timeconsuming procodure. Endofree plasmid kits, provide optimal transfection results for ordering information, see pages 3336. Qiaamp dna mini and blood mini handbook moodle ufsc. Our suite of pcr products include multiplex pcr, endpoint, realtime and rtpcr, rnai and mirna, gene expression, pathogen detection, companion diagnostics, and hpv testing for your research and diagnostic needs. Plasmid mini kits can be used with any microcentrifuge or. The objective of this experiment is to introduce the principles of extracting plasmid dna from bacterial cells. We initially tried two different commercial miniprep kits.
These are available online in convenient and compact pdf format at where you can find, view, and print the msds for each qiagen kit and kit component. These columns use a unique silica membrane to achieve high yields of up to 40 g of sequencinggrade plasmid dna. Qiagen plasmid mini handbook 0399 9 protocol plasmid mini purification protocol this protocol is for mini up to 20 g preparations of highcopy plasmid dna from cultures of e. Dna purification from dried blood spots qiaamp dna mini kit. Product use limitations the rneasy plus mini kit is intended for molecular biology applications. Purifying plasmid dna using the qiagen miniprep kit protocol. Product information thermo scientific genejet plasmid. Add 10 ml of buffer p3, mix gently, and incubate on ice for 1520 min.
Principle qiaamp viral rna mini kits provide the fastest and easiest way to purify viral rna for reliable use in amplification technologies. Quickstart protocol march 2016 qiagen plasmid mini, midi and. As part of the stringent qiagen quality assurance program, the performance of qiaprep miniprep kits is monitored routinely on a lottolot basis. For purification of lowcopy plasmids and cosmids, large plasmids 10. C and is stable for at least 9 months under these conditions. Bac prep using quiagen plasmid mini prep kit do not need. If you have any questions or experience any difficulties regarding qiaamp dna mini or qiaamp dna blood mini kits, or qiagen products in general, please do not hesitate. Plasmid dna prepared by tianprep mini plasmid kit is suitable for a variety of routine applications including restriction enzyme digestion, sequencing, library screening, ligation and trans. Qiagen plasmid kits are intended for molecular biology applications. For the fastest, most convenient purification of up to 10 mg transfectiongrade plasmid dna. One such kit which is often used is from the qiagen company. Quickstart protocol march 2016 qiagen plasmid mini, midi. The objective of this lab is to perform extraction of plasmid dna and analyze the results. Optimized protocols for specific sample types provide reproducible purification of highquality dna for life science, genotyping, and veterinary pathogen research applications.
Simply add the uniquely formulated 7x lysis buffer directly to your bacterial culture, neutral. Features of the purelink quick plasmid miniprep kit. The zyppy plasmid miniprep kit is a plasmid dna kit that features a pelletfree modified alkaline lysis system that bypasses the normal cellpelleting and resuspension steps common to classical plasmid preparation procedures. These are available online in convenient and compact pdf. Simply add the uniquely formulated 7x lysis buffer directly to. Plasmid mini kit, supplied by qiagen, used in various techniques. The dna yield of qiaprep miniprep kits is tested by preparing highcopy. By combining silicabinding technology and the convenience of a spin column format, up to 15. Plasmid dna encoding constitutively expressed gfp pegfpc2 was prepared using either monarch plasmid miniprep kit or qiagen qiaprep spin miniprep kit. Lysis by adding 250 ul solution p2, sit at room temp 5 min 5. Maxipreps for plasmid dna purificationnucleobond xtra maxi. All due care and attention should be exercised in the handling of the products. Objectives after completion, the student should be able to.
1247 136 726 667 53 568 1399 1175 1433 437 219 1234 922 1296 1331 80 148 1567 1374 1779 1134 1365 288