Cambio - Excellence in Molecular Biology

Enzymes for Molecular Biology

Enzymes for Molecular Biology: Novel Enzymes (New for 2019)

New range of enzymes for 2019. Our new range of enzymes include novel enzymes specifically designed or engineered to be amazingly tolerant to high salt concentrations, function at very low temperatures and heat inactivated.

Cryophile Shrimp Alkaline Phosphatase (heat-labile)

Cryophile Shrimp Alkaline Phosphatase (heat-labile / heat-inactivatable) is a recombinant enzyme that originates from the arctic shrimp (Pandalus borealis).

Cambio

Catalogue No.DescriptionPack SizePriceQty
CA-1736-1000Cryophile Shrimp Alkaline Phosphatase (heat-labile)1000 U (1U/µl) £80.00 Quantity Add to Order
CA-1736-5000Cryophile Shrimp Alkaline Phosphatase (heat-labile)5000 U (1U/µl) £310.00 Quantity Add to Order
CA-1736-5000GFCryophile Shrimp Alkaline Phosphatase (heat-labile)5000U (>20U/µl) Glycerol free POA Quantity Add to Order

Cryophile Shrimp Alkaline Phosphatase (heat-labile)

Cryophile Shrimp Alkaline Phosphatase (heat-labile / heat-inactivatable) is a recombinant enzyme that originates from the arctic shrimp (Pandalus borealis).

Cambio

Cryophile Shrimp Alkaline Phosphatase (heat-labile) is a recombinant enzyme that originates from the arctic shrimp (Pandalus borealis). Given its heat-labile properties, Cambio's Shrimp Cryophile Alkaline Phosphatase is more suitable for many applications than other native phosphatases (E. coli or calf intestine).

The Cryophile Shrimp Alkaline Phosphatase has become one of today’s best-selling DNA modifying enzymes due to the added convenience of complete heat inactivation. While most other alkaline phosphatases (from E. coli or calf intestine) must be removed by extraction procedures, Cryophile Shrimp Alkaline Phosphatase is completely inactivated after 5 minutes at 65°C.  It works well in common buffers without the requirement for other additions.

Main advantages with Cryophile Shrimp Alkaline Phosphatase

  • Very high specific activity
  • 100% heat-inactivatable at 65°C
  • Removes phosphates from DNA, RNA, dNTPs and dephosphorylates proteins
  • May be added directly to restriction enzyme digests
  • No vector purification necessary
  • Requires no supplemental zinc or other additives for activity
  • Works directly in most relevent buffers
  • Easy treatment of unincorporated dNTPs in PCR products prior to DNA sequencing or SNP analysis

See also details of Dephosphorylation in the applications tab above

 

 

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200

Cryophile Shrimp Alkaline Phosphatase (heat-labile)

Cryophile Shrimp Alkaline Phosphatase (heat-labile / heat-inactivatable) is a recombinant enzyme that originates from the arctic shrimp (Pandalus borealis).

Cambio

Dephosphorylation prior to cloning

In cloning protocols, a DNA fragment is ligated into a plasmid vector. If the vector is cut with a single restriction enzyme, the chances are much higher that the vector re-ligates back on itself rather than on an added DNA fragment. This results in a high fraction of “empty clones”, or background.

Alkaline phosphatases are routinely used to reduce the background from empty, re-ligated vectors during cloning of DNA fragments, since dephosphorylated DNA termini cannot be ligated by DNA ligase. The phosphatase treatment will effectively reduce the background of “empty” clones by >95%. However, cloning procedures and also phosphatase treatment may be cumbersome and error-prone.

Cryophile Shrimp Alkaline Phosphatase offers greater convenience for this procedure, since the enzyme activity may be completely removed by a simple heat inactivation step. Cryophile Shrimp Alkaline Phosphatase is active in all buffers used for restriction enzymes, so can be added either during restriction digestion, or directly after. With Cryophile Shrimp Alkaline Phosphatase the user can forget elaborate calculations and multi-step incubations, because the enzyme completely dephosphorylate DNA during one, simple, incubation.

There are many ways to use Cryophile Shrimp Alkaline Phosphatase for vector dephosphorylation, and we recommend two protocols below. These protocols apply to all types of DNA termini: 3’-protruding, blunt, or 5’-protruding.

Protocol including restriction cutting

In this protocol, Cryophile Shrimp Alkaline Phosphatase is present during restriction cutting, so the termini are dephosphorylated as soon as they are formed. In this protocol, the minimum effective amount of Cryophile Shrimp Alkaline Phosphatase is proportional to the amount of restriction enzyme added (i.e. the rate of terminus formation). In the simple sense, use at least 0.1U of Cryophile Alkaline Phosphatase per unit of restriction enzyme, and proceed to complete cutting. The amount of restriction enzyme may differ from the protocol below; please use amounts recommended.

  • 1 µg plasmid
  • 5 Unit restriction enzyme
  • 5 µl 10x restriction enzyme buffer
  • 1 Unit Cryophile Shrimp Alkaline Phosphatase
  • dH2O to 50 µl

Incubate at 37°C for 1 hour, inactivate as recommended for the restriction enzyme used. Proceed to ligation protocol.

Quick dephosphorylation of cut plasmid

Efficient dephosphorylation can be achieved in short time using high amounts of Cryophile Shrimp Alkaline Phosphatase. When restriction cutting is complete, simply add 5 U Cryophile Shrimp Alkaline Phosphatase per µg vector to your restriction mix and incubate for further 5 min at 37°C. Inactivate as recommended for the restriction enzyme. Proceed to ligation protocol.

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200