Product Specifications R4160F Rev A
Product Description: HaeIII is a DNA endonuclease that
recognizes the double-strand DNA sequence below and cuts
the phosphodiester backbone in both strands at the dotted
line.
Product Information
HaeIII
Part Number
R4160F
Concentration
10,000 U/mL
Unit Size
5’ GG⁞CC 3’
3’ CC⁞GG 5’
4,000 U
Storage Temperature
-25°C to - 15°C
Lot Number
Reference Number
Units Tested
SDS
Purity
n/a
SS
Exonuclease
100 U
DS
Exonuclease
100 U
Specification
>95%
< 1.0%
Released
<1.0%
Released
Assay
Product Specifications
R4160 F
E. coli DNA
Contamination
50 U
RNase
Contamination
100 U
Phosphatase
Contamination
100 U
End Integrity (Ligation
and Recutiting)
20 per digest
<10 copies
Not Detected
Not Detected
> 95% ligation, > 95%
of ligation is recut
Source of Protein: A recombinant Haemophilus aegytius strain carrying the HaeIII gene
Unit Definition: One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 37°C in a total
reaction volume of 50 µl.
Molecular weight: 37,128 Daltons
Quality Control Analysis:
Unit Activity is measured using a 2-fold serial dilution method. Dilutions of enzyme were made in 10X Emerald Reaction Buffer
and added to 50 μL reactions containing 1 μg of λ DNA. Reactions are incubated for 1 hour at 37°C, stopped, and analyzed on a
0.8% agarose gel stained with ethidium bromide.
Protein Concentration (OD280) is determined by OD280 absorbance.
Physical Purity is evaluated by SDS-PAGE of concentrated and diluted enzyme solutions followed by silver stain detection.
Purity is assessed by comparing the aggregate mass of contaminant bands in the concentrated sample to the mass of the
protein of interest band in the diluted sample.
Single-Stranded Exonuclease is determined in a 50 µL reaction containing a radiolabeled single-stranded DNA substrate and 10
µL of enzyme solution incubated for 4 hours at 37°C.
Double-Stranded Exonuclease is determined in a 50 µl reaction containing a radiolabeled double-stranded DNA substrate and
10 µL of enzyme solution incubated for 4 hours at 37°C.
E.coli 16S rDNA Contamination is evaluated using 5 µL replicate samples of enzyme solution denatured and screened in a
TaqMan qPCR assay for the presence of contaminating E.coli genomic DNA using oligonucleotide primers corresponding to the
16S rRNA locus.
Product Specifications R4160F Rev (A)
Non-Specific RNAse contamination is assessed using the RNAse Alert kit, (Integrated DNA Technologies), following the
manufacturer’s guidelines.
Phosphatase Contamination is determined by measuring the conversion of PNPP to p-nitrophenol (A405nm) after 4 hour
incubation at 37⁰C.
End integrity (ligation and re-cutting) following over-digestion of DNA substrate with restriction enzyme, >95% of the DNA
fragments can be ligated with T4 DNA ligase in 16 hours at 16°C. Of the ligated fragments >95% can be recut with restriction
endonuclease.
Supplied in:
10 mM Tris-HCl, 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 200 µg/mL BSA, 50% Glycerol pH 7.4 @ 25°C
Notes:
Requires Mg2+ for activity
dam and dcm methylation: Not Sensitive
CpG methylation: Not Sensitive
Heat Inactivation: 80°C for 20 minutes
Excess enzyme, extended incubation times, use of non-optimal buffers and/or excess glycerol (>10% v/v) may cause the
enzyme to cut at sites other than its recognition site.
Limitations of Use
This product was developed, manufactured, and sold for in vitro use only. The product is not suitable for administration to humans or
animals. MSDS sheets relevant to this product are available upon request.
100 Cummings Center, Suite 407J, Beverly, MA 01915 • Ph (888) 927-7027 • Fax (978) 867-5724 • www.enzymatics.com FMWI016.1 Rev D