ATAC-Seq试剂盒

在全基因组范围内分析开放的染色质区域。

 

ATAC-Seq试剂盒总览

ATAC-Seq是一种快速检测方法,可通过识别具有开放或可及状态的染色质区域来分析整个基因组的表观遗传概况。ATAC-Seq方法快速,简便,适用于广泛的样品类型,已成为常用的表观遗传学分析方法,并且可以作为更深入的表观遗传学分析的基础。

在ATAC-Seq实验中,用高活性的突变Tn5转座酶处理来自细胞或组织样品的完整细胞核。该转座酶把DNA片段化的同时用测序接头标记靶DNA。

ATAC-Seq Kit

Active Motif的ATAC-Seq试剂盒提供了与Illumina®平台兼容的ATAC-Seq文库制备试剂,可以从20 – 30 mg组织或50,000 – 100,000个细胞中生成16种不同的ATAC-Seq文库。经过优化的方案将指导您完成样品制备、DNA片段标记和文库构建的步骤,构建可在单个测序反应中进行多重测序的文库。

ATAC-Seq试剂盒亮点

  • 显示开放染色质区域的基因组序列
  • 仅需数小时即可完成NGS文库制备
  • 简单,快速的实验步骤与优化的试剂
了解有关ATACT-Seq的更多信息
 

ATAC-Seq试剂盒组分

ATAC-Seq试剂盒包含两盒组分,其中-20°C保存的一盒组分干冰运输,4°C保存的另一盒组分室温运输。首次使用后,请根据说明书中的存放条件保存。妥善保管的试剂自收货之日起可稳定保存6个月。

试剂包括

  • ATAC Lysis Buffer, store at RT
  • Assembled Transposomes, store at -20°C
  • 2X Tagmentation Buffer, store at -20°C
  • 1X PBS, store at -20°C or RT
  • 10X PBS, store at RT
  • 10% Tween 20, store at RT
  • 0.5% Digitonin, store at RT
  • DNA Purification Columns, store at RT
  • DNA Purification Binding Buffer, store at RT
  • DNA Purification Wash Buffer, store at RT
  • DNA Purification Elution Buffer, store at RT
  • 3 M Sodium Acetate, store at RT
  • 10 mM dNTPs, store at -20°C
  • 5X Q5 Buffer, store at -20°C
  • Q5 High-Fidelity DNA Polymerase, store at -20°C
  • i7 Indexed Primer 1, store at -20°C
  • i7 Indexed Primer 2, store at -20°C
  • i7 Indexed Primer 3, store at -20°C
  • i7 Indexed Primer 4, store at -20°C
  • i5 Indexed Primer 1, store at -20°C
  • i5 Indexed Primer 2, store at -20°C
  • i5 Indexed Primer 3, store at -20°C
  • i5 Indexed Primer 4, store at -20°C
  • SPRI Beads, store at 4°C
 

ATAC-Seq试剂盒数据

ATAC-Seq试剂盒包含进行16个ATAC-Seq反应所需的试剂和酶。这些试剂盒组分经过优化,可以高效工作,经验证可以生成高质量的ATAC-Seq数据。

揭示启动子处染色质的开放位置

具有转录活性的基因,其启动子处的染色质通常是开放的或可及的。ATAC-Seq分析会在这些位置产生峰,揭示正在研究的样品中开放的染色质区域。

ATAC-Seq Kit Genome Browser

图1:ATAC-Seq试剂盒实验结果展示在基因组浏览器中。ATAC-Seq试剂盒用于评估小鼠脑组织样品的全基因组开放染色质状态。此处显示的基因组浏览器轨迹位于FOXP3基因位置,该基因编码有丝分裂后神经元的生物标记物。峰表明在FOXP3基因的启动子上有开放的染色质。

试剂盒实验结果与已发表数据高度相关

Active Motif的ATAC-Seq试剂盒可生成高质量数据。使用ATAC-Seq试剂盒产生的ATAC-Seq数据与已发表的ATAC-Seq数据具有很好的相关性,这突出说明了该检测方法的可靠性和可重复性。

ATAC-Seq RKPM Data

图2:ATAC-Seq试剂盒文库与已发表的ATAC-Seq数据集的相关性。该图显示了在GM12878细胞中,已发表的ATAC-Seq(Buenrostro, J. D. et al. (2013) Nat. Methods 10: 1213-1218)数据的峰中的RPKM信号与使用试剂盒生成的数据集之间的相关性为0.94。

 

ATAC-Seq Kit FAQs

Is this kit the same as the original Buenrostro et al Nature 2013 paper?

This kit is based off a combination of the original Buenrostro paper and the Corces et al Nature 2017 paper, and we have optimized the buffers for a more robust and efficient assay.

What is the source of the recombinant Tn5 enzyme in this kit?

The recombinant Tn5 enzyme in this kit is a hyperactive mutant that we cloned, expressed, purified, and test in-house. The Assembled Transposomes containing this enzyme are provided pre-loaded with the next-gen sequencing adapters, so it is ready-to-use without any optimization or other preparation required.

What is the minimum number of cells or tissue per reaction?

50,000 cells or 20 mg tissue.

Can frozen tissue and cells be used?

Yes, but the samples must be high quality to preserve viability. Cells should be cryopreserved in a controlled-rate freeze with media formulated to protect against ice crystal formation and subsequent cell damage, and tissue should be flash frozen at -80°C.

Can nuclei be frozen for later use in ATAC-Seq?

Yes, extracted nuclei can be cryopreserved and frozen at -80°C for later use. Nuclei can also be pooled together to increase the sample size.

Can FACS-sorted cells be used?

It is possible, but cells could be damaged during the FACS-sorting process. For good ATAC-Seq libraries, cells must be viable.

Do you have a recommendation for a 40 micron mesh strainer for tissue samples?

Yes, we recommend the Falcon 40 micron cell strainer (Falcon catalog no. 352340).

Is DNase treatment recommended for all samples?

DNase treatment can help in certain situations such as mitochondrial DNA depletion, or if there are 15% dead cells in the sample, but it can end up removing cells you want data from if the cells aren’t healthy and able to exclude the enzyme. It’s only recommended as an option when cells are viable and won’t also be digested by the enzyme.

I don’t have a thermomixer. Can I still use the kit?

Using a thermomixer is recommended. However, we have tested this and incubated the tagmentation reaction without the use of a mixer at 37°C in a thermal cycler with a heated lid and the reaction was successful.

Can all the reagents and kit components, including master mixes and enzymes, thaw out and be prepared at room temperature?

Everything except the Assembled Transposomes and Q5 DNA polymerase can be thawed at room temperature. The Assembled Transposomes and Q5 DNA Polymerase are formulated with glycerol and will not need thawing, just put these on ice. The other components can be frozen again and thawed again for future reactions, so not all 16 reactions need to be done simultaneously.

What is the expected final library size, and what should the library traces look like on a fragment analyzer?

ATAC-Seq Kit Library Traces

(Click image to enlarge size)

Library fragments should range from around 250 bp to 1000 bp in length with an oscillation period ~150 bp.

How many sequencing reads are needed per sample?

Typically 20M reads.


 

ATAC-Seq试剂盒文档

 

 

 

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