Duplex-specific nuclease

Duplex-specific nuclease

cat.# EA001; EA002; EA003

- Selectively cleaves ds DNA and DNA in DNA-RNA hybrid duplexes
- Discriminates between perfectly and nonperfectly matched short duplexes
- Inactive towards ss DNA and RNA
- Thermostable
- Inhibited by EDTA

Duplex-specific nuclease (DSN) is an enzyme purified from hepatopancreas of Red King (Kamchatka) crab (Shagin et al., 2002). DSN shows a strong preference for cleaving double-stranded (ds) DNA and DNA in DNA-RNA hybrid duplexes, and is practically inactive towards single-stranded (ss) DNA or single or double-stranded RNA. Moreover, the enzyme is capable to discriminate between perfectly and nonperfectly matched short DNA duplexes. DSN is widely used for full-length enriched cDNA normalization (Zhulidov et al., 2004; Bogdanova et al., 2011a) and for construction of normalized RNA-seq libraries for next generation sequencing ((Christodoulou et al., 2011), Illumina DSN normalization protocol). The list of approved DSN applications also includes normalization of genomic DNA (Shagina et al., 2010), cDNA depletion and ribosomal cDNA depletion (Bogdanova et al., 2009; Bogdanova et al., 2011b; Yi et al., 2011), cDNA subtraction (Peng et al., 2008), SNP detection (Shagin et al., 2002; Liu et al., 2011), construction of repeat-free FISH probes (Swennenhuis et al., 2012), multiplexed fluorescence detection of miRNAs (Yin et al., 2012), quantitative telomeric overhang determination (Zhao et al., 2008; Zhao et al., 2011), etc.

Available products

PRODUCTCAT.#DESCRIPTIONSIZEPRICEUSER MANUAL (PDF)
The prices do not include delivery. The prices vary in different countries. Please contact your local distributor for exact prices and delivery information.
Duplex-specific nuclease, lyophilized EA001 Thermostable nuclease from Red King (Kamchatka) crab hepatopancreas with a strong preference for cleaving double-stranded DNA and DNA in DNA-RNA hybrid duplexes 50 U € 350
EA002 100 U € 500
EA003 10 U € 130

DSN was purified from Red King (Kamchatka) crab hepatopancreas using modified protocol from Shagin et al., 2002. DNAase activity was measured using modified Kunitz assay (Kunitz, 1950), where unit definition was defined as: the amount of DSN added to 50 μg/ml calf thymus DNA that causes an increase of 0.001 absorbance units per minute. Activity assay was performed at 25°C, in 50 mM Tris-HCl buffer, pH 7.15, containing 5 mM MgCl2.

Shipping and Storage conditions:
All components of the kit can be shipped at ambient temperature. Upon arrival all components must be stored at -20°C.

Notice to Purchaser:

DSN-related materials (also referred to as "Products") are intended for research use only. The Products are covered by U.S. Patents No. 7,435,794 and 7,803,922. By use of these Products, you accept the terms and conditions of the applicable Limited Use Label License #002.

References:

  • Bogdanova EA, Barsova EV, Shagina IA, Scheglov A, Anisimova V, Vagner LL, Lukyanov SA, Shagin DA. Normalization of full-length-enriched cDNA. Methods Mol Biol. 2011a; 729 :85-98. doi: 10.1007/978-1-61779-065-2_6 / pmid: 21365485
  • Bogdanova EA, Shagina IA, Mudrik E, Ivanov I, Amon P, Vagner LL, Lukyanov SA, Shagin DA. DSN depletion is a simple method to remove selected transcripts from cDNA populations. Mol Biotechnol. 2009; 41 ((3)):247-53. doi: 10.1007/s12033-008-9131-y / pmid: 19127453
  • Bogdanova EA, Shagina IA, Yanushevich YG, Vagner LL, Lukyanov SA, Shagin DA. Preparation of prokaryotic cDNA for full-scale transcriptome analysis. Russian Journal of Bioorganic Chemistry. 2011b; 37 (6):775-8. ISI:000297344600012 http://www.springerlink.com
  • Christodoulou DC, Gorham JM, Herman DS, Seidman JG. Construction of normalized RNA-seq libraries for next-generation sequencing using the crab duplex-specific nuclease. Curr Protoc Mol Biol. 2011; Chapter 4 :Unit4.12. doi: 10.1002/0471142727.mb0412s94 / pmid: 21472699
  • Kunitz M. Crystalline desoxyribonuclease; digestion of thymus nucleic acid; the kinetics of the reaction. J Gen Physiol. 1950; 33 :363-377. / pmid: 15406374
  • Liu M, Yuan M, Lou X, Mao H, Zheng D, Zou R, Zou N, Tang X, Zhao J. Label-free optical detection of single-base mismatches by the combination of nuclease and gold nanoparticles. Biosens Bioelectron. 2011; 26 (11):4294-300. doi: 10.1016/j.bios.2011.04.014 / pmid: 21605966
  • Peng RH, Xiong AS, Xue Y, Li X, Liu JG, Cai B, Yao QH. Kamchatka crab duplex-specific nuclease-mediated transcriptome subtraction method for identifying long cDNAs of differentially expressed genes. Anal Biochem. 2008; 372 (2):148-55. / pmid: 17905189
  • Shagin DA, Rebrikov DV, Kozhemyako VB, Altshuler IM, Shcheglov AS, Zhulidov PA, Bogdanova EA, Staroverov DB, Rasskazov VA, Lukyanov S. A novel method for SNP detection using a new duplex-specific nuclease from crab hepatopancreas. Genome Res. 2002; 12 (12):1935-42. / pmid: 12466298
  • Shagina I, Bogdanova E, Mamedov IZ, Lebedev Y, Lukyanov S, Shagin D. Normalization of genomic DNA using duplex-specific nuclease. Biotechniques. 2010; 48 (6):455-9. doi: 10.2144/000113422 / pmid: 20569220
  • Swennenhuis JF, Foulk B, Coumans FA, Terstappen LW. Construction of repeat-free fluorescence in situ hybridization probes. Nucleic Acids Res. 2012; 40 (3):e20. doi: 10.1093/nar/gkr1123 / pmid: 22123742
  • Yi H, Cho YJ, Won S, Lee JE, Jin Yu H, Kim S, Schroth GP, Luo S, Chun J. Duplex-specific nuclease efficiently removes rRNA for prokaryotic RNA-seq. Nucleic Acids Res. 2011; 39 (20):e140. doi: 10.1093/nar/gkr617 / pmid: 21880599
  • Yin BC, Liu YQ, Ye BC. One-step, multiplexed fluorescence detection of microRNAs based on duplex-specific nuclease signal amplification. J Am Chem Soc. 2012; 134 (11):5064-7. doi: 10.1021/ja300721s / pmid: 22394262
  • Zhao Y, Hoshiyama H, Shay JW, Wright WE. Quantitative telomeric overhang determination using a double-strand specific nuclease. Nucleic Acids Res. 2008; 36 (3):e14. / pmid: 18073199
  • Zhao Y, Shay JW, Wright WE. Telomere G-overhang length measurement method 1: the DSN method. Methods Mol Biol. 2011; 735 :47-54. doi: 10.1007/978-1-61779-092-8_5 / pmid: 21461810
  • Zhulidov PA, Bogdanova EA, Shcheglov AS, Vagner LL, Khaspekov GL, Kozhemyako VB, Matz MV, Meleshkevitch E, Moroz LL, Lukyanov SA, Shagin DA. Simple cDNA normalization using kamchatka crab duplex-specific nuclease. Nucleic Acids Res. 2004; 32 (3):e37. / pmid: 14973331
Copyright 2002-2016 Evrogen. All rights reserved.
Evrogen JSC, 16/10 Miklukho-Maklaya str., Moscow, Russia, Tel +7(495)988-4084, Fax +7(495)988-4085, e-mail:evrogen@evrogen.com