商品编号


H0221S



品牌


Biohelix



公司


BIO-HELIX Co., LTD.



公司分类


Primer Navigator? series



Size

50?g


Concentration

0.5 mg/ml
















商品信息



Description:


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Single-Strand Binding Protein (SSB) binds to single-stranded DNA with high affinity and also binds to RNA and double-stranded DNA with lower affinity (1).
In vivo
, it st
ABI
lizes transiently formed ssDNA and plays an important role in DNA replication, recombination and repair (2).
In vitro
, SSB proteins have been used to dest
ABI
lize secondary structures in DNA and to increase the processivity of DNA polymerases in several molecular
BIOLOG
y applications: SSBs improve the yield and efficiency of reverse transcription reactions during RT-PCR as well as increase the yield of PCR products (3-9).

ET SSB (Extreme
Thermo
stable SSB)
is a single-stranded DNA binding protein isolated from a hyper
Thermo
philic microorganism, and it is a flagship enhancer of the Primer Navigator product series. It remains fully active after incubation at 95?C for 60 min. Due to the extreme
Thermo
st
ABI
lity,
ET SSB
can be used in applications that require extremely high temperature conditions, such as nucleic acid amplification and sequencing.




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Source:


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Purified from an
E. coli
strain that overexpresses the ssb gene isolated from a hyper
Thermo
philic microorganism.



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Applications:


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Improve the yield of multiplex PCR (Fig.1) and multiplex HDA (Fig.2)

Improve the processivity of DNA polymerase (10)

St
ABI
lization and marking of ssDNA structure (11)

Increase the yield and specificity of PCR reactions (5-9)

Increase the yield and processivity of RT during RT-PCR (3,4)

Improve DNA sequencing through regions with strong secondary structure (8)

Enhance the RecA activity for ssDNA binding and strand trasfer (12,13)




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Storage Conditions:


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Concentration:

0.5mg/ml
Storage Buffer:

20 mM Tris-HCl
200 mM NaCl
1 mM EDTA
0.5 mM DTT
50% Glycerol
pH7.5 @ RT

Storage Temperature:
-20?C




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Quality Control:


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Quality Assurance Statement:

ET SSB is purified free of contaminating endonucleases and exonucleases. Each lot is tested for single-strand, DNA-dependent ATPase activity and is visually determined to be > 95% pure on an SDS-polyacrylamide gel.

Exonuclease Activity:

Incubation of 20 ?g ET SSB for 4 hours at 37°C in 50 ?l reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 ?g of a mixture of single and double-stranded [
3
H]
E. coli
DNA (200,000 cpm/?g) released < 0.05% of the total r
ADI
oactivity.

Endonuclease Assay:

Incubation of 10 ?g ET SSB for 4 hours at 37°C in 50 ?l reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 ?g ΦX174 RF I DNA gave < 5% conversion to RF II.

Nuclease Activity:

Incubation of 20 ?g ET SSB for 16 hours at 37°C in 50 ?l of reaction buffer containing 50 mM potassium acetate, 20 mM Tris-acetate, 10 mM magnesium acetate and 1 mM dithiothreitol, pH 7.9 @ 25°C, with 1 ?g λ DNA yielded a clear and sharp band on an agarose gel.




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References:


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Coleman, J. E. & Oakley, J. L. (1980) CRC Crit Rev Biochem 7, 247-89.

Chase, J. W. & Williams, K. R. (1986) Annu Rev Biochem 55, 103-36.

Baugh, L. R., Hill, A. A., Brown, E. L. & Hunter, C. P. (2001) Nucleic Acids Res 29, E29.

Villalva, C., Touriol, C., Seurat, P., Trempat, P., Delsol, G. & Brousset, P. (2001) Biotechniques 31, 81-3, 86.

Schwarz, K., Hansen-Hagge, T. & Bartram, C. (1990) Nucleic Acids Res 18, 1079.

Chou, Q. (1992) Nucleic Acids Res 20, 4371.

Oshima, R. G. (1992) Biotechniques 13, 188.

Rapley, R. (1994) Mol Biotechnol 2, 295-8.

Olszewski, M., Rebala, K., Szczerkowska, Z. & Kur, J. (2005) Mol Cell Probes 19, 203-5.

Myers, T. W. & Romano, L. J. (1988) J Biol Chem 263, 17006-15.

Delius, H., Mantell, N. J. & Alberts, B. (1972) J Mol Biol 67, 341-50.

Reddy, M. S., Vaze, M. B., Madhusudan, K. & Muniyappa, K. (2000) Biochemistry 39, 14250-62.

West, S. C., Cassuto, E. & Howard-Flanders, P. (1982) Mol Gen Genet 186, 333-8.



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