T4 Endonuclease VII
T4 Endonuclease VII

Endonuclease VII is the product of gene 49 of bacteriophage T4. It has a mass of 18 kDa. T4 Endonuclease VII involves in DNA-packaging, genetic recombination and mismatch repair in vivo.
 
Name Cat# Size Concentration Price Qty
T4 Endonuclease VII
ENDO7-100
2,500 U
10,000 U/ml
$210.00
T4 Endonuclease VII
ENDO7-200
10,000 U
10,000 U/ml
$630.00
T4 Endonuclease VII
ENDO7-OEM
please inquire

Description:
Endonuclease VII is the product of gene 49 of bacteriophage T4. It has a mass of 18 kDa. T4 Endonuclease VII involves in DNA-packaging, genetic recombination and mismatch repair in vivo. It has also been demonstrated in vitro to resolve single-base misparings, heteroduplex loops and branched DNAs, such as four-way Holliday junctions and three-way Y-structures. 

CRISPR-Cas9 System

Source:
A recombinant E. coli strain carrying the cloned T4 Endonuclease VII gene
 
Unit Definition:
One unit (0.5 ng) of the enzyme resolves 50% of 1 pmol FAM labeled 28mer oligonucleotide substrate [1] within the immobile 4-way Holliday junction structure in 30 minutes at 37°C in 50mM Tris-HCl, pH 8.0, 10mM MgCl2, 10mM 2-ME and 0.1 μg/μl BSA. 

Specific Activity:  2000 U/µg

Recommended Storage Condition: -20°C

Experimental Data:
T4 Endonuclease VIIT4 Endonuclease VII

Figure1.  Performance of MCLAB’s T4 Endonuclease VII analyzed by capillary electrophoresis. (a) Negative control sample analysis, 10 pmol of FAM labeled 28mer oligonucleotide substrate. (b) 40 U of MCLAB’s T4 Endonuclease VII was able to fully resolve 10 pmol of FAM labeled 28mer oligonucleotide substrate with a 4-way Holliday junction structure (30 minutes at 37°C in 50 mM Tris-HCl, pH 8.0, 10 mM MgCl2, 10 mM 2-ME and 0.1 μg/μl BSA).
T4 Endonuclease VII
Figure2.  MCLAB’s T4 Endonuclease VII and T7 Endonuclease I (from other supplier) activity comparison.  (a) 10 U (275 fmol) of MCLAB T4’s Endonuclease VII can resolve 64% of 10 pmol of 4-way junction substrate (enzyme to substrate molar ratio 1:70) in 30 minutes at 37°C in 50 mM Tris-HCl, pH 8.0, 10 mM MgCl2, 10 mM 2-ME and 0.1 μg/μl BSA. (b) 40 U (368 fmol) of T7 Endonuclease I (supplier N) only resolves 43% of 10 pmol of 4-way junction substrate (enzyme to substrate molar ratio 1:50) in 30 minutes at 37°C in 50 mM NaCl, 10 mM Tris-HCl, pH 7.9, 10 mM MgCl2, 1 mM DTT. Compared to T7 Endonuclease I (supplier N), MCLAB’s T4 Endonuclease VII resolves 4-way Holliday Junctions at a higher yield in 30 minutes with less enzyme.
T4 Endonuclease VII
T4 Endonuclease VII Protocol

For 4-way junction substrate:

1.       Prepare the following reaction:

Component

Volume

Water

16 µl

10x Endo VII reaction buffer

2 µl

10µM 4-way Junction (FAM labeled)

1 µl

Endo VII (dilute if necessary)

1 µl

Total 

20 µl

 

2.       Mix well and incubate the reaction @ 37°C for 30min.

3.       Use 1 µl reaction to analyze the cleaved fragments on Capillary electrophoresis.           

 

For single base mismatch substrate:

1.       Use about 200-400ng DNA fragment that contains single base mismatch for each reaction:

Component

Volume

10x Endo VII buffer

1 µl

Substrate

200-400ng

Endo VII enzyme (dilute if necessary)

1 µl

Water

To total volume of 10 µl

Total 

10 µl

 

2.      Mix well and incubate the reaction @ 37°C for 30min.

3.      Run a 2% agarose gel and check for cleaved bands. 

 

Table. Comparison of T4 Endonuclease VII and T7 Endonuclease I

 

T4 Endonuclease VII

T7 Endonuclease I

Protein Mass

18 KDa

60 kDa

Function

Resolvase

Resolvase

Application

Enzymatic mutation detection
Resolve branched DNA
Detect or cleave heteroduplexes

Enzymatic mutation detection
Resolve branched DNA
Detect or cleave heteroduplexes

Source

Bacteriophage T4

Bacteriophage T7

Protein Design

T4 Endonuclease VII

A fusion of maltose binding protein (MBP) and T7 Endonuclease I

Activity
(Tested with 4-way Holliday junctions in 30 minutes.)

High*
10 units of the enzyme (278 fmol protein, molar ratio 1:70 (Enzyme:Substrate)) resolve 64% of 4-way Junctions.

Low*
40 units of the enzyme (368 fmol protein, molar ratio 1:50 (Enzyme:Substrate)) resolve 43% of 4-way Junctions.

Specificity

High* (single resolved peak shown on CE)

Low* (multiple resolved peaks shown on CE)

* See Figure 2 for T4 endonuclease VII and T7 endonuclease I activity comparison result.

Supplied in:
10 mM Tris-HCl
50 mM KCl 
1 mM DTT
0.1 mM EDTA 
50% Glycerol 
pH 7.4 @ 25°C

Supplied With:
10x T4 Endonuclease VII Reaction Buffer:
500 mM Tris-HCl
100 mM MgCl2
100 mM 2-mercaptoethanol
1 mg/ml BSA
pH 8.0 @ 25°C

Reference:
1. Golz, S., Birkenbihl, R. P., and Kemper, B. (1995), DNA Research 2, 277-284.

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