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You are here:Home » Molecular Biology » MB-Enzymes, Restriction » Csp6I (RsaI)

Csp6I (RsaI)

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Specifications

5'-G^T A C-3'
3'-C A T^G-5'
Unlike RsaI, Csp6I produces DNA fragments with a 2-base 5'-extension.
Catalog #C7946
SourceCorynebacterium species RFL6
Concentration10u/ul
Unit DefinitionOne unit is defined as the amount of enzyme required to digest 1ug of lambda DNA in 1 hour at 37°C in 50ul of assay buffer.
Storage BufferSupplied as a liquid in 10mM Tris-HCl, pH 7.4 at 25°C, 100mM potassium chloride, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA, 50% glycerol.
Supplied WithR1625 Restriction Enzyme Buffer A, 10X: Supplied as a liquid in33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA, pH 7.9 at 37°C.
R1625-01 Restriction Enzyme Buffer B, 10X Supplied as a liquid in 10mM Tris-HCl, pH 7.5, 10mM magnesium chloride, 0.1mg/ml BSA. Incubate at 37°C
Enzyme Properties:
Stability during Prolonged Incubation:
A minimum of 0.1u of Csp6I is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal Inactivation: Csp6I is inactivated by incubation at 65°C for 20min.
Compatible Ends: FspBI, NdeI, Tru1I, VspI
Number of Recognition Sites in DNA:
Lambda: 113
PhiX174: 11
pBR322, pUC57, pUC18/19: 3
pTZ19R/U: 2
M13mp18/19: 19
Methylation Effects:
Dam, Dcm, EcoKI, EcoBI: Never overlaps-no effect
CpG: May overlap-no effect
Quality Control:
Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 320-fold overdigestion (20u/ug lambda DNA x 16 hours) with Csp6I.
Ligation/Recutting Assay: After 50-fold overdigestion (3u/ug DNA x 17 hours) with Csp6I more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 05uM. More than 95% of these can be recut.
Labeled Oligonucleotide Assay: No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10u of Csp6I for 4 hours.
Protocol for Digestion
Add
Nuclease free water16ul
R1625-012ul
DNA (0.5-1ug/ml)1ul
C79460.5-2ul
Mix gently and spin down for a few seconds. Incubate at 37ºC for 1-16 hours.
Protocol for Digestion Directly after Amplification
Add
PCR Reaction Mixture10ul (~0.1-0.5ug of DNA)
Nuclease free water18ul
R1625-012ul
C79461-2ul
Mix gently and spin down for a few seconds.
Incubate at 37ºC for 1-16 hours.
Storage and Stability
May be stored at 4°C for short-term only. For long-term storage, store at -20°C. Aliquots are stable for at least 12 months at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.


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