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 | | | | | NEBNext® Quick DNA Sample Prep Reagent Set 2 |
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Description:
The NEBNext® Quick DNA Sample Prep Reagent Set 2 contains enzymes and buffers that are ideally suited for sample preparation for next-generation sequencing, and for preparation of expression libraries. Each of these components must pass rigorous quality control standards and are lot controlled, both individually and as a set of reagents.
Lot Control: The lots provided in the Quick DNA Sample Prep Reagent Set 2 are managed separately and are qualified by additional functional validation. Individual reagents undergo standard enzyme activity and quality control assays, and also meet stringent criteria in the additional quality controls listed on each individual component page.
Functionally Validated: Each set of reagents is functionally validated together through construction and sequencing of a genomic DNA library on a Roche 454 GS FLX Titanium™ System, and by construction of an expression library.
For larger volume requirements, customized and bulk packaging is available by purchasing through the OEM/Bulks department at NEB. Please contact OEM@neb.com for further information.
The Reagent Set Includes:
The volumes provided are sufficient for preparation of up to 10 reactions (NEB #E6080S) and 50 reactions (NEB #E6080L). (All reagents should be stored at -20°C).
NEBuffer 2 (10X)
#E6081A: 0.025 ml
#E6081AA: 0.125 ml
1X NEBuffer 2:
10 mM Tris-HCl
50 mM NaCl
10 mM MgCl21 mM DTT
pH 7.9 @ 25°C
Deoxynucleotide Solution Mix (10 mM each dNTP)
#E6083A: 0.015 ml
#E6083AA: 0.075 ml
Description: Deoxynucleotide Solution Mix is an equimolar solution of ultrapure dATP, dCTP, dGTP and dTTP, provide for the PCR enrichment reaction.
Supplied in: Milli-Q™ water as a sodium salt at pH 7.5.
Concentration: Each nucleotide is supplied at a concentration of 10 mM. (40 mM total nucleotide concentration).
Quality Assurance: Nucleotide solutions are certified free of nucleases and phosphatases.
Notes: Storing nucleotide triphosphates
T4 DNA Polymerase
#E6084A: 0.015 ml
#E6084AA: 0.075 ml
Description: T4 DNA Polymerase catalyzes the synthesis of DNA in the 5´→ 3´ direction and requires the presence of template and primer. This enzyme has a 3´→ 5´ exonuclease activity which is much more active than that found in DNA Polymerase I. Unlike E. coli DNA Polymerase I, T4 DNA Polymerase does not have a 5´→ 3´ exonuclease function.
Source: Purified from a strain of E. coli that carries a T4 DNA Polymerase overproducing plasmid.
Supplied in: 100 mM KP04 (pH 6.5), 1 mM DTT and 50% glycerol.
Adenosine 5´- Triphosphate (10 mM)
#E6082A: 0.025 ml
#E6082AA: 0.125 ml
Description: Adenosine 5´-Triphosphate (ATP) is a substrate for ATP-dependent enzyme systems.
Supplied in: Sterile purified water adjusted to pH 7.0 with NaOH
Molecular Weight: 551.2 daltons (disodium salt)
T4 Polynucleotide Kinase
#E6085A: 0.015 ml
#E6085AA: 0.075 ml
Description: Catalyzes the transfer and exchange of Pi from the γ position of ATP to the 5´-hydroxyl terminus of polynucleotides (double- and single-stranded DNA and RNA) and nucleoside 3´-monophosphates. Polynucleotide Kinase also catalyzes the removal of 3´-phosphoryl groups from 3´-phosphoryl polynucleotides, deoxynucleoside 3´-monophosphates and deoxynucleoside 3´-diphosphates (1).
Source: An E. coli strain that carries the cloned T4 Polynucleotide Kinase gene. T4 Polynucleotide Kinase is purified by a modification of the method of Richardson (1).
Supplied in: 50 mM KCl, 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM dithiothreitol, 0.1 μM ATP and 50% glycerol.
Quality Assurance: Free of exonuclease, phosphatase, endonuclease and RNase activities. Each lot is tested under 5´-end-labeling conditions to assure maximal transfer of [32P].
Taq DNA Polymerase
#E6086A: 0.015 ml
#E6086AA: 0.075 ml
Description: Taq DNA Polymerase is a thermostable DNA polymerase that possesses a 5´→3´ polymerase activity and a double-strand specific 5´→3´ exonuclease activity.
Source: Purified from an E. coli strain that carries the Taq DNA polymerase gene from Thermus aquaticus YT-1.
Supplied in: 10 mM Tris-HCl, 100 mM KCl, 1 mM Dithiothreitol, 0.1 mM EDTA, 50% Glycerol, 0.5% Tween-20 and 0.5% NP-40, pH 7.4 @ 25°C.
Quick T4 DNA Ligase
#E6087A: 0.015 ml
#E6087AA: 0.075 ml
Source: Purified from E. coli C600 pcl857 pPLc28 lig8 (2).
NEBNext Sizing Buffer (1.2X)
#E6088A: 15 ml
#E6088AA: 75 ml
1X NEBNext Sizing Buffer:
7% Polyethylene Glycol 8000
1 M NaCl
Description: NEBNext Sizing Buffer has been optimized for use with AMPure beads to select DNA fragments > 300 bp from a sample of mixed sized DNA fragments.
TE Buffer (1X)
#E6089A: 5 ml
#E6089AA: 25 ml
Description: TE Buffer is free of detectable DNA and RNA nucleases and is suitable for us in molecular biology applications.
Required Materials Not Included:
Agencourt® Ampure® Beads (Beckman Coulter)
DNA Adaptors or Vectors
Magnetic Separation Rack (NEB #S1510)
Storage Conditions
Storage Temperature:
-20°C
Protocols
Protocols for NEBNext® Quick DNA Sample Prep Reagent Set 2
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Quality Control Assays:
Please refer to the Instruction Manual for details.
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