QIAGEN Large-Construct Kit
For purification of up to 50 μg BAC, PAC, and P1 DNA or up to 200 μg cosmid DNA, free of genomic DNA
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QIAGEN Large-Construct Kit (10)
Cat no. / ID. 12462
10 QIAGEN-tip 500, Reagents, Buffers, ATP-Dependent Exonuclease
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This product contains substances regulated under REACH (EC 1907/2006 Annex XIV). The use of this product in the EU is permitted subject to an exemption (Article 56(3)). Please refer to the REACH notification and the SDS of this product, both of which can be found in the “Resources” section of this page, for more information.
The QIAGEN Large-Construct Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
✓ 24/7 automatic processing of online orders
✓ Knowledgeable and professional Product & Technical Support
✓ Fast and reliable (re)-ordering
Features
- Efficient removal of genomic DNA contamination
- Pure, transfection-grade large-construct DNA
- Easy and simple procedure
Product Details
The QIAGEN Large-Construct Kit provides gravity-flow, anion-exchange columns for purification of large-molecular-weight DNA. A unique integrated ATP-dependent exonuclease digestion step ensures selective removal of contaminating genomic DNA. The purified DNA is equivalent to that obtained by 2 x CsCl gradient centrifugation and is suitable for transfection.
Performance
DNA purified using the QIAGEN Large-Construct Kit is free of genomic DNA contamination (see figure " Efficient removal of genomic DNA"). This removal of genomic DNA ensures accurate and reliable DNA quantitation, which enables sensitive experiments to be carried out under defined and reproducible conditions.
See figures
Principle
DNA purification using the QIAGEN Large-Construct Kit uses an optimized gravity-flow procedure which yields DNA of significantly greater purity than that obtained with other commonly used methods. A unique integrated treatment with ATP-Dependent Exonuclease enables efficient removal of genomic DNA.
The unique anion-exchange resin in QIAGEN-tips, included in the QIAGEN Large-Construct Kit, is developed exclusively for the purification of nucleic acids. Its exceptional separation properties result in DNA purity equivalent or superior to that obtained by two successive rounds of CsCl gradient centrifugation. Prepacked QIAGEN-tips (see figure " Anion-exchange tips") operate by gravity flow and never run dry, minimizing the hands-on time required for plasmid preparation. The entire QIAGEN plasmid purification system avoids the use of toxic substances such as phenol, chloroform, ethidium bromide, and CsCl, minimizing hazard both to the user and the environment.
See figures
Procedure
Following alkaline lysis of up to 500 ml culture (see flowchart " QIAGEN Plasmid Kit procedures"), a unique integrated digestion step with ATP-dependent exonuclease provided with the kit, ensures selective removal of contaminating genomic DNA, as well as nicked or damaged construct DNA. The sample is then loaded onto the anion-exchange tip, where plasmid DNA selectively binds under appropriate low-salt and pH conditions. RNA, proteins, metabolites, and other low-molecular-weight impurities are removed by a medium-salt wash. Genomic DNA-free, pure plasmid DNA is eluted in high-salt buffer. The DNA is concentrated and desalted by isopropanol precipitation and collected by centrifugation.
See figures
Applications
DNA purified using the QIAGEN Large-Construct Kit is suitable for any application, including:
- Subcloning and preparation of shotgun libraries
- Direct sequencing of large-construct DNA
- Transfection
Supporting data and figures
QIAGEN Plasmid Kit procedures.
Neutralized bacterial lysates are cleared by centrifugation. The cleared lysate is then loaded onto the anion-exchange tip where plasmid DNA selectively binds under appropriate low-salt and pH conditions. RNA, proteins, metabolites, and other low-molecular-weight impurities are removed by a medium-salt wash, and ultrapure plasmid DNA is eluted in high-salt buffer. The DNA is concentrated and desalted by isopropanol precipitation and collected by centrifugation.
Specifications
| Features | Specifications |
|---|---|
| plasmidtype | BAC, PAC, P1, cosmid DNA |
| applications | Subcloning, transfection, sequencing etc. |
| processing | Manual (centrifugation) |
| culturevolumestartingmaterial | <500 ml culture volume |
| samplesperrunthroughput | 1 sample per run |
| technology | Anion-exchange technology |
| timeperrunorprepperrun | 280 min |
| yield | <150 ug |
Resources
Quick-Start Protocols (2)
Kit Handbooks (1)
Safety Data Sheets (1)
Certificates of Analysis (1)
Publications
Targeted disruption of Kaposi's sarcoma-associated herpesvirus ORF57 in the viral genome is detrimental for the expression of ORF59, K8alpha, and K8.1 and the production of infectious virus.
J Virol; 2006; 81 (3):1062-71 2006 Nov 15 PMID:17108026
Functional characterization of Kaposi's sarcoma-associated herpesvirus ORF45 by bacterial artificial chromosome-based mutagenesis.
J Virol; 2006; 80 (24):12187-96 2006 Oct 11 PMID:17035322
Autographa californica multiple nucleopolyhedrovirus nucleocapsid assembly is interrupted upon deletion of the 38K gene.
J Virol; 2006; 80 (23):11475-85 2006 Sep 20 PMID:16987976
Identification of in vivo-induced bacterial protein antigens during human infection with Salmonella enterica serovar Typhi.
Infect Immun; 2006; 74 (9):5161-8 2006 Sep PMID:16926408
Molecular characterization of a diagnostic DNA marker for domesticated tetraploid wheat provides evidence for gene flow from wild tetraploid wheat to hexaploid wheat.
Mol Biol Evol; 2006; 23 (7):1386-96 2006 May 4 PMID:16675504