Recombinant S. marcescens NucA Protein, CF

Compare to Benzonase® Nuclease
Catalog # Availability Size / Price Qty
10038-NA-020
10038-NA-100
NucA Protein Enzyme Activity
1 Image
Product Details
FAQs
Reviews

Recombinant S. marcescens NucA Protein, CF Summary

Purity
>90%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Activity
Measured by its ability to hydrolyze DNA from salmon testes. The specific activity is >130,000 pmol/min/ug, as measured under the described conditions.
Source
E. coli-derived s. marcescens NucA protein
Met1-Asn266
Accession #
Predicted Molecular Mass
28.9 kDa
SDS-PAGE
29 kDa, reducing conditions

Product Datasheets

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

10038-NA

Formulation Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Shipping The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Assay Buffer: 100 mM Tris, 1 mM MgCl2, pH 8.5
  • rS.marcescens NucA (Catalog # 10038-NA)
  • Deoxyribonucleic acid (DNA) sodium salt from salmon testes (Millipore Sigma, Catalog # D1626), 5 mg/mL stock in deionized water
  • Coupling Enzyme: Recombinant Mouse Alkaline Phosphatase/ALPL (rmALPL) (Catalog # 2910-AP)
  • Malachite Green Phosphate Detection Kit (Catalog # DY996)
  • 96-well Clear Plate (Catalog # DY990)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute 1 M Phosphate Standard by adding 10 µL of the 1 M Phosphate Standard to 990 µL of Assay Buffer for a 10 mM stock. Continue by adding 10 µL of the 10 mM Phosphate stock to 990 µL of Assay Buffer for a 100 µM stock. This is the first point of the standard curve.
  2. Complete the standard curve by performing six one-half serial dilutions of the 100 µM Phosphate stock in Assay Buffer. The standard curve has a range of 0.078 to 5 nmol per well.
  3. Prepare a reaction mixture containing 0.16 mg/mL DNA and 4 µg/mL rmALPL in Assay Buffer.
  4. Dilute rS. marcescens NucA to 4 ng/mL in Assay Buffer.
  5. Load 50 µL of each dilution of the standard curve into a plate. Include a curve blank containing 50 μL of Assay Buffer.
  6. Load 25 µL of 4 ng/mL rS. marcescens NucA into empty wells of the same plate as the curve. Include a Control containing 25 µL of Assay Buffer.
  7. Add 25 µL of reaction mixture to the wells, excluding the standard curve.
  8. Seal plate and incubate at room temperature for 20 minutes.
  9. Add 30 µL of the Malachite Green Reagent A to all wells. Mix briefly.
  10. Add 100 µL of deionized water to all wells. Mix briefly.
  11. Add 30 µL of the Malachite Green Reagent B to all wells. Mix and incubate for 20 minutes at room temperature.
  12. Read plate at 620 nm (absorbance) in endpoint mode.
  13. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Phosphate released* (nmol) x (1000 pmol/nmol)
Incubation time (min) x amount of enzyme (µg)

  *Derived from the phosphate standard curve using linear or 4-parameter fitting and adjusted for Control.

Per Reaction:
  • rS. marcescens NucA: 0.1 ng (0.0001 µg)
  • rmALPL: 0.1 µg
  • DNA: 4 µg

Data Image

Enzyme Activity NucA (Benzonase)Digestion of DNA.  View Larger

2 mg plasmid DNA was incubated with increasing amounts(ng/mL) of Recombinant S. marcescens NucA (Benzonase) (Catalog # 10038-NA) in50 mM Tris (pH 8.0) and 1 mM MgCl2 for 30 minutes at room temperature.Samples were analyzed by agarose gel electrophoresis and DNA was visualized byethidium bromide staining.

Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

=
÷

Background: NucA

Serratia marcescens endonuclease is a non-specific nuclease that degrades both single- and double-stranded nucleic acids, including DNA and RNA, but show no proteolytic activity (1); therefore, it is ideal for the removal of nucleic acid contaminants from protein samples and applications where complete digestion of nucleic acids is desirable. The optimum pH for enzyme activity is found to be 8.0-9.2. It hydrolyzes internal phosphodiester bonds between nucleotides in nucleic acids to produce 5'-monophosphate oligonucleotides of 3-8 bases in length (2). The active enzyme is a homodimer with two disulfide bonds in each monomer that are crucial to the enzyme activity and stability (3). The absolute activity of the recombinant enzyme is measured by a phosphatase coupled assay (4), where the 5'-phosphate of oligosaccharides generated by the enzyme is further released by non-specific alkaline phosphatase and quantitated by Malachite reagents (5).

References
  1. Benedik, MJ and Strych, U. (1998) FEMS Microbiol Lett. 165:1.
  2. Nestle, M, et al. (1999) J. Biol. Chem. 274:825.
  3. Ball, T.K. et al. (1992) Nucleic Acids Res. 20:4971.
  4. Wu, Z.L. et al. (2011) Glycobiology 21:727.
  5. Van Veldhoven, P.P. and G.P. Mannaerts (1987) Anal. Biochem. 161:45.
Long Name
Nuclease A
Alternate Names
NucA

FAQs

No product specific FAQs exist for this product, however you may

View all Proteins and Enzyme FAQs

Reviews for Recombinant S. marcescens NucA Protein, CF

There are currently no reviews for this product. Be the first to review Recombinant S. marcescens NucA Protein, CF and earn rewards!

Have you used Recombinant S. marcescens NucA Protein, CF?

Submit a review and receive an Amazon gift card.

$25/€18/£15/$25CAN/¥75 Yuan/¥1250 Yen for a review with an image

$10/€7/£6/$10 CAD/¥70 Yuan/¥1110 Yen for a review without an image

Submit a Review