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작성일 : 12-08-14
[RiboSolutions] Benzonase and Cyanase as probes for chromatin accessibility
Interesting method for genome mapping of chromatin using Benzonase and Cyanase versus DNase
Benzonase and Cyanase as probes for chromatin accessibility
 
 
 
Background
 
The challenge in extracting genome-wide chromatin features from limiting clinical samples poses a significant hurdle in identification of regulatory marks that impact the physiological or pathological state. Current methods that identify nuclease accessible chromatin are reliant on large amounts of purified nuclei as starting material. This complicates analysis of trace clinical tissue samples that are often stored frozen. We have developed an alternative nuclease based procedure to bypass nuclear preparation to interrogate nuclease accessible regions in frozen tissue samples.
 
 
Results
 
Here we introduce a novel technique that specifically identifies Tissue Accessible Chromatin (TACh). The TACh method uses pulverized frozen tissue as starting material and employs one of the two robust endonucleases, Benzonase or Cyansase, which are fully active under a range of stringent conditions such as high levels of detergent and DTT. As a proof of principle we applied TACh to frozen mouse liver tissue. Combined with massive parallel sequencing TACh identifies accessible regions that are associated with euchromatic features and accessibility at transcriptional start sites correlates positively with levels of gene transcription. Accessible chromatin identified by TACh overlaps to a large extend with accessible chromatin identified by DNase I using nuclei purified from freshly isolated liver tissue as starting material. The similarities are most pronounced at highly accessible regions, whereas identification of less accessible regions tends to be more divergence between nucleases. Interestingly, we show that some of the differences between DNase I and Benzonase relate to their intrinsic sequence biases and accordingly accessibility of CpG islands is probed more efficiently using TACh.
 
 
Conclusion
 
The TACh methodology identifies accessible chromatin derived from frozen tissue samples. We propose that this simple, robust approach can be applied across a broad range of clinically relevant samples to allow demarcation of regulatory elements of considerable prognostic significance.
 
 
관련 article 전문(PDF)
 
(미국 국립보건원에 의해 소개된 자료입니다. / Publication date: 26 June 2012)
 
 
Benzonase Alternative Cyanase Nuclease System
 
 
Cyanase Nuclease는 박테리아 유래의 높은 활성을 가진 핵산 분해 효소입니다. 다양한 종류의 DNA 및 RNA를 비특이적으로 분해할 수 있습니다. 또한 Cyanase Nuclease와 높은 affinity를 가진 Cyanase Inactivation Resin에 의해, 반응 후의 Cyanase Nuclease를 반응 용액에서 제거할 수 있습니다. 단백질 등의 시료에서 핵산 제거에 유용합니다.
 
 
Features
  • Cell Lysate Clearance, Protein and Viral Purification, 시료 등 에서의 핵산 제거에 최적입니다.
  • Degrades all forms of DNA and RNA
  • Easily inactivated and removed
  • Fastest Nuclease on the Market. Bar none
  • Unsurpassed stability. Up to 8 months at room temperature. No other competitor comes close.
 
핵산 제거 효소로 범용되고있는 Serratia marcescens 유래의 neclease보다 cost performance가 높고, 대체품으로 사용하실 수 있습니다. pH와 MgSO 4 등 폭넓은 조성 조건에서도 활성을 가지고 있습니다 (아래 그림 참조).
Lysozyme, DTT(<100 mM), 계면 활성제(Triton X-100과 Tween 20을 포함)에도 영향을 받지 않습니다.
 
  
 
3 μg의 λDNA를 이용하여 다양한 조성 조건에서 Cyanase 의한 분해를 실시했다. 반응은 0.5 U 또는 1.0 U의 효소와 버퍼를 이용해 모두 37 ℃에서 1 분간 행하였다.
 
Lane 1 : 1.0 kb 마커,
Lane 2 : 제품 0.5 U
Lane 3 : 제품, 1.0 U
Lane 4 : Serratia marcescens 유래의 Nuclease 0.5 U
Lane 5 : Serratia marcescens 유래의 Nuclease 1.0 U
Lane 6 : DNase, 0.5 U
Lane 7 : DNase, 1.0 U
Lane 8 : 1.0 kb 마커
 
 
 
Ordering information ;
 
10,000 U와 25,000 U의 2가지 kits가 있습니다.
 
 
 
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