Pkr I

Source: Planomicrobium koreense 78k

 

 

 

 

 

 

 

1000 u/ml               store at -20°C

 

Recognition sequence

 

GCN↑GC

CG↓NCG

 

Source:  Planomicrobium koreense 78k

 

The enzyme cleaves C5-methylated DNA and doesn't cut unmodified DNA.

 

PkrI cleaves DNA sequence 5`- GCNGC-3`/3`-CGNCG-5', if at least three 5-methylcytosines are present in the recognition site (N isn`t considering) [1].

 

Optimal recognition sites (100% activity):

5`-G(5mC)NG(5mC)-3`/3`(5mC)GN(5mC)G-5`

5`-G(5mC)NGC-3`/3`-(5mC)GN(5mC)G-5`

5`-GCNG(5mC)-3`/3`-(5mC)GN(5mC)G-5`

   

Supplied in:

20 mM Tris-HCl (pH 7.4); 200 mM NaCl; 0,1 mM EDTA; 7 mM  2-mercaptoethanol; 50% glycerol.

 

Reaction Conditions:

1X SEBuffer Y. 

 

Incubate at  37°C.

 

1X SEBuffer Y  (pH 7.9 @ 25ºC)

33 mM Tris-Ac, 66 mM KAc, 10 mM MgAc, 1 mM DTT

 

Unit Definition:

One unit is defined as the amount of enzyme required to hydrolyze at least one of three canonical sites: 5`-G(5mC)NG(5mC)-3`/3`-CGN(5mC)G-5` in 1 μg of linearized plasmid pFsp4HI3 in 1 hour at 37°C in a total reaction volume of 50 μl. DNA pFsp4HI3/DriI is a linearized plasmid pFsp4HI3, which carries a gene of DNA-methyltransferase M.Fsp4HI and includes  three canonical sites: 5`-G(5mC)NG(5mC)-3`/3`-CGN(5mC)G-5` [2].

 

Quality Control Assays

 

16-Hour Incubation:

No detectable degradation of 1μg of Lambda DNA was observed after incubation with 2 units of enzyme for 16 hours at 37°C in a total reaction volume of 50 μl.

 

Oligonucleotide Assay:

No detectable degradation of a single- and double-stranded oligonucleotide was observed after incubation with 2 units of enzyme for 3 hours.

 

Enzyme Properties

 

Activity in SEBuffers:

SEBuffer B      50-75%

SEBuffer G     75-100%

SEBuffer O      10-25%

SEBuffer W     25-50%

SEBuffer Y         100%

SEBuffer ROSE  100%

When using a buffer other than the optimal (suppied) SEBuffer, it may be necessary to add more enzyme to achieve a complete digestion.

 

Reagents supplied with enzyme:

10X SEBuffer Y

 

Heat Inactivation:

Yes   (65°C for 20 minutes)

 

Pkr I

 Restriction Endnuclease from Sibenzyme

 

* availibility of sample size may be limited

96,00 €
order / kaufen
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  • 1 - 3 Tage Lieferzeit / 1-3 days delivery1

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Datasheet

Material Safety Datasheet


References / Protocols / Notes / Recomendations / Tests

 

 

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