SNPase for SNP-Genotyping

Hot-Start Polymerase for SNP detection by allele-specific PCR and micro sequencing

Cat.-no	Description	Amount	Price €	Shop
S500	SNpase	500 units	80.00	add
S505	SNpase	5x500 units	349.00	add

SNPase for SNP detection: Datasheet
SNPase for SNP detection: Deutsche Beschreibung

Features:
- 10-15 fold lower mutation rate than Taq DNA Polymerase
- high fidelity allele-specific amplification of DNA fragments
- high specificity with lowest background AS-PEX and AS-PCR
- Hot-Start activity for less primer dimers
- only 5'-3' polymerase activity, lack of 5’-exonuclease activity

Applications:
- High specific PCR
- Multiplex PCR
- Real-Time PCR with intercalation dyes
- high fidelity dNTPs and ddNTPs
- Mini-Sequencing, SNP-genotyping

Description:
SNPase is Taq DNA Polymerase with unique N-terminal deletion and proprietary amino acids substitutions introduced into the active center of the enzyme. This modification causes dramatic increase of sensitivity of the enzyme to mismatches at 3’-end of the primer. Consequently , non-perfect annealing of the primers does not result in unspecific amplicons formation. This enzyme has only 5'-3' polymerase activity and is recommended for SNP genotyping by allele-specific PCR (AS-PCR), allele-specific primer extension (AS-PEX) and minisequencing procedures.
Minisequencing SNP genotyping with SNPase DNA Polymerase can be carried out by the procedure
described in *:
* Reference for minisequencing protocol: Lovmar L, Fredriksson M, Liljedahl U, Sigurdsson S, Syvänen AC.
Quantitative evaluation by minisequencing and microarrays reveals accurate multiplexed SNP genotyping of whole genome amplified DNA. Nucleic Acids Res. 2003;31:e129.

Unit definition:
One unit is defined as the amount of enzyme that incorporates 10nmoles of dNTPs into acid-insoluble
form in 30 minutes at 72°C.

Concentration:
20-25 u/µl

Reaction Buffer supplied:
5X Reaction buffer without MgCl₂MgCl₂ 100 mM

Note:
- optimal MgCl₂ concentration: 3.0 -3.5 mM in the 1X reaction mixture
- higher MgCl₂ concentrations results in higher yield (up to 4.5 mM)
- lower MgCl₂ (2.5 mM) results in higher specificity
- DNA fragments up to 400 bp from Human genomic DNA and 500 bp from Phage-DNA

Usage:

Components Volume per reaction

5X reaction buffer 5 µl

MgCl₂ 2.5 - 4 mM

dNTP-Mix 0.2 mM each

primer mix (5 µM stock) 0,9-1,1 µl (5 pmol)

Template DNA 75-125 ng/25 µl genomic DNA

SNpase 0.2 - 0.5 µl (5-12 units)

Sterile dest. Water (molecular grade) up to 25 µl total reaction volume

General Thermo-Cycler protocol:

Step Time Temperature

Initial denaturation 1-2 min 94-95°C

30-35 Cycles:
Denaturation
Annealing
Extension
20-30 sec
15-30 sec
30-40 sec
94-95°C
59-68°C
68-72°C per 1kb

Final extension 5 min 72°C