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商品編號


L6060L



品牌


Enzymatics



公司


Enzymatics,inc



公司分類(lèi)


Ultrapure Ligases



Size

20,000 U

商品信息

Taq DNA Ligase


Product Description

Taq
DNA Ligase catalyzes the formation of a phosphodiester bond in duplex DNA containing adjacent 5′-phosphoryl and 3′-hydroxyl termini, using NAD
+
as a cofactor.

Source of Protein
A recombinant
E. coli
strain carrying the cloned
Taq
DNA Ligase gene.

Supplied in
10 mM Tris-HCl
50 mM KCl
1 mM DTT
0.1 mM EDTA
0.1% Tween-20
50% glycerol
pH 7.5 @ 25°C

Supplied With
B6060 (10X
Taq
DNA Ligase Buffer)

10X
Taq
DNA Ligase Buffer (B6060)
200 mM Tris-HCI
250 mM KCl
100 mM MgCl
2
5 mM NAD
0.1% Triton X-100
pH 7.6 @ 25°C

Unit Definition
1 unit is defined as the amount of
Taq
DNA Ligase required to join 50% of 1 ?g of the 12-base cohesive ends of Lam
BD
a DNA cut with Sma I and Sal I in 50 ?l 1X
Taq
DNA Ligase Buffer following a 10 minute incubation at 45°C.



Quality Control Analysis

Unit Characterization Assay
Unit activity was measured using a 2-fold serial dilution method. Dilutions of enzyme batch were made in 1X
Taq
DNA Ligase Reaction Buffer and added to 50 ?L reactions containing λ Hind III digested DNA and 1X Taq DNA Ligase Reaction Buffer. Reactions are incubated for 10 minutes at 45°C, stopped, and analyzed on a 0.8% agarose gel stained with ethidium bromide.

SDS-Page (Physical Purity Assessment)
2.0 ?L of concentrated enzyme solution was loaded on a denaturing 4-20% Tris-Glycine SDS-PAGE gel flanked by a broad-range MW
Marker
and 2.0 ?L of a 1:100 dilution of the same enzyme species. Following electrophoresis, the gel was stained and the samples compared to determine physical purity. The acceptance criteria for this test requires that the aggregate mass of contaminant bands in the concentrated sample do not exceed the mass of the protein of interest band in the dilute sample, confirming greater than 99% purity of the concentrated sample.



Contamination Tests

Single-Stranded Exonuclease Activity
A 50 ?L reaction containing 10,000 cpm of a r
ADI
olabeled single-stranded DNA substrate and 10 ?L of enzyme solution incubated for 4 hours at 37°C resulted in less than 5.0% release of TCA-soluble counts.

Double-Stranded Exonuclease Activity
A 50 ?l reaction containing 5,000 cpm of a r
ADI
olabeled double-stranded DNA substrate and 10 ?L of enzyme solution incubated for 4 hours at 37°C resulted in less than 1.0% release of TCA-soluble counts.

Double-Stranded Endonuclease Activity
A 50 ?L reaction containing 0.5 ?g of pENZuC DNA and 10 ?L of enzyme solution incubated for 4 hours at 37°C resulted in no visually discern
IBL
e conversion to nicked circular DNA as determined by agarose gel electrophoresis.

E.coli
16S rDNA Contamination test
Replicate 5 ?L samples of enzyme solution were 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. The acceptance criterion for the test is the threshold cycle count (C
t
) produced by the average of 3 replicate no template control samples. Based on the correlation between the no template control C
t
values and standard curve data, the detection limit of this assay is <10 copies genome/sample.


View PDF Poster Instructions FAQ


Product Information



Taq DNA Ligase


Part Number
L6060L
Price
$363
Concentration
40,000 U/ml
Unit Size
20,000 U


SDS

Available on request








Product Specification*


Storage Temperature
-25 to -15°C


Test

Specification


Purity (SDS-PAGE)
>99%
Specific Activity
400,000 U/mg
SS Exonuclease
400 U <5.0% released
DS Exonuclease
400 U <1.0% released
DS Endonuclease
400 U = No conversion
E.coli
DNA Contamination
400 U <10 copies





* For a detailed summary of assay conditions and data, refer to the Quality Controls Analysis section.

Legal Disclaimer

Certain applications in which this product can be used may be covered by patents issued and applicable in the United States and abroad. Purchase of this product does not include a license to perform any patented application; therefore it is the sole responsibility of users of this product to determine whether they may be required to engage a license agreement depending upon the particular application in which the product is used.



Limitations of Use
This product was developed, manufactured, and sold for in vitro use only. The product is not suitable for admi
NIST
ration to humans or animals. SDS sheets relevant to this product are available upon request.