对多种起始材料进行DNA亚硫酸氢盐转化,无需单独的DNA纯化步骤
  • 适用于基因组DNA、全血、细胞、组织和FFPE组织切片
  • 高效进行样本裂解和亚硫酸氢盐转化
  • 产物为即用型DNA,产量高,洗脱体积小
  • 独特的DNA保护技术可防止DNA降解
  • 最大程度上减少手工操作,快速获得结果

EpiTect Plus Bisulfite Kits是一组整合的试剂盒体系,适用于各种起始材料,可高效的进行样本裂解,直接对从全血、培养细胞、组织样本和FFPE组织切片中提取的DNA进行亚硫酸氢盐转化。转化后的DNA可洗脱到10 µl的洗脱缓冲液中,因此DNA的浓度高。EpiTect Plus Bisulfite Kits还具有同样高的DNA转化效率,以及EpiTect Bisulfite Kits可靠的DNA保护技术。

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EpiTect Plus DNA Bisulfite Kit
48 MinElute DNA spin columns, Bisulfite mix, DNA Protect Buffer, Carrier RNA, Buffers
59124
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EpiTect Plus Bisulfite Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.


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DNA storage stability.|DNA Protect Buffer.|Superior yields.|High recovery of converted DNA.|EpiTect Plus DNA flowchart.|EpiTect Plus LyseAll flowchart.|EpiTect Plus FFPE flowchart.|Interconnected protocols.|
Bisulfite converted DNA generated with EpiTect Kits was stored at –20°C for up to 36 months, and amplified at several time intervals. The resulting threshold cycle values at all time points for various DNA amounts were comparable to those obtained with freshly converted DNA (time point 0).|[A] Upon addition to the bisulfite conversion reaction, the DNA Protect Buffer [B] should turn from green to blue, indicating sufficient mixing and correct pH.|DNA was purified from rat liver FFPE tissue (10 µm) using the EpiTect Plus FFPE Bisulfite Kit (1–4) or an alternative kit from Supplier Z (5–8). [A] Detection of the indicated genes using the PyroMark PCR Kit and agarose gel analysis. [B] Detection of two amplicons of the GSTP gene (156 and 279 bp) using the QuantiTect SYBR® Green PCR Kit (4 triplicate samples). Templates purified using the EpiTect Plus FFPE Bisulfite Kit resulted in lower CT values and highly reproducible results for both amplicons (left). Templates purified using the kit from Supplier Z resulted in amplification of only the 156 bp fragment with higher CT values (right).|Degradation of DNA during the bisulfite conversion reaction results in low yields of the target sequence for downstream applications. This problem is particularly important for samples such as FFPE tissues, where the starting DNA is already fragmented. The DNA Protect Buffer included in EpiTect Plus Bisulfite Kits minimizes DNA damage during conversion. Along with minimal loss of DNA during sample lysis and thorough cleanup of the resulting converted DNA, this innovative DNA protect technology maximizes recovery of the converted DNA.|Streamlined and straightforward, the EpiTect Plus DNA Bisulfite Kit enables complete bisulfite conversion and clean up of genomic DNA in approximately 6 hours.|Upon completion of the sample lysis steps, bisulfite conversion is completed using the included EpiTect Plus DNA Bisulfite Kit.|Upon completing deparaffinization, lysis, and decrosslinking steps, bisulfite conversion is completed using the EpiTect Plus DNA Bisulfite Kit.|Depending on sample type, the bisulfite conversion of DNA calls for the use of a specific kit and protocol. EpiTect Plus Bisulfite Kits contain buffers optimized for DNA extraction and all reagnets for the DNA bisulfite conversion.|
Performance
使用EpiTect Plus Bisulfite Kits得到的DNA的产量是其他试剂盒的4–8倍(参见"High recovery of converted DNA"和"Superior yields")。产量增加,使得下游操作更加灵活;因为由一个样本得到的转化后DNA可用于多个后续操作,或储存以备将来使用。目前QIAGEN的数据显示:使用EpiTect Plus Bisulfite Kits转化和纯化的DNA可稳定储存至少3年(参见"DNA storage stability")。
Principle

EpiTect Plus Bisulfite Kits的主要特点包括:

DNA的转化效率超过99%,且DNA的降解程度达到最小
转化后的DNA浓度高,洗脱体积小(低至10 µl)
整合的技术可最大程度上减少样本裂解过程中的DNA损失和保护DNA,转化效率高
优化、简便的操作流程适用于基因组DNA、全血、培养细胞、组织样本和FFPE组织切片
起始材料用量灵活:1 ng–2 µg gDNA、0.5–20 µl全血、10–100,000个细胞或1片FFPE组织切片(10 µm厚)
可进行组织裂解,节省时间

使用亚硫酸氢钠进行DNA转化时,所有样本DNA中非甲基化的胞嘧啶都被完全转化,同时减少了由于温度和化学试剂因素导致的DNA降解。对于转化后DNA的保护十分重要,影响回收率和下游操作的灵敏度。EpiTect Plus Bisulfite Kits在防止DNA降解的同时,可高效的进行DNA亚硫酸氢盐转化,且无需单独的DNA纯化步骤。这个高效的工作流程覆盖了从样本裂解到最终洗脱转化后DNA的整个过程,可快速获得适合多种下游应用的高品质、即用型DNA。

QIAGEN对DNA亚硫酸氢盐转化的解决方案—找到适合您的试剂盒

3种EpiTect Plus Bisulfite Kits相互关联,是一个整体,这3种试剂盒可对多种起始材料的DNA进行直接的亚硫酸氢盐转化(参见"Interconnected protocols")。但每种试剂盒有自己独特的特点,适用于某一特定样本类型,含有进行特定转化所需的所有缓冲液和试剂。使用每个试剂盒所含的试剂可进行高效的样本裂解,裂解过程的DNA损失降到最小;完全的亚硫酸氢盐转化,转化过程中有效的保护DNA和彻底回收DNA。

EpiTect Plus Bisulfite Kits不能替代EpiTect Bisulfite Kits,但适用于更多样本类型,并可为下游应用制备亚硫酸氢盐转化DNA。两套试剂盒都有DNA保护技术,可确保高转化效率。EpiTect Bisulfite Kits提供便利的96孔板,适合高通量转化。下表"QIAGEN对DNA亚硫酸氢盐转化的解决方案"中列出了QIAGEN表观遗传学产品线中的5个亚硫酸氢盐转化试剂盒,以便您从中选择合适的试剂盒。

整合的技术—DNA Protect Buffer和Deparaffinization Solution

独特配方的DNA Protect Buffer,可预防在高温、低pH环境下对DNA进行亚硫酸氢盐处理时产生的DNA片段化。该缓冲液可促使DNA变性,生成胞嘧啶完全转化所需的单链DNA。DNA Protect Buffer含有pH指示染料,可确保胞嘧啶转化时pH准确(参见"DNA Protect Buffer")。

脱蜡过程可将DNA从FFPE样本中提取出来。在该过程中,石蜡先被溶解,然后去除;再用裂解缓冲液和蛋白酶K处理样本。Deparaffinization Solution是新型试剂,可减少去石蜡化过程所需的手动操作,并可确保高DNA回收率。在溶液中加入FFPE样本后,溶液作用于样本,同时蛋白酶K开始消化。之后无需离心沉淀FFPE样本,也无需去除含有石蜡的上清。因此避免了去石蜡化过程中的样本损失。

QIAGEN对DNA亚硫酸氢盐转化的解决方案
特点 EpiTect Bisulfite Kit EpiTect 96 Bisulfite Kit EpiTect Plus DNA Bisulfite Kit EpiTect Plus LyseAll Bisulfite Kit  EpiTect Plus FFPE Bisulfite Kit
规格 EpiTect离心柱 96孔板 MinElute DNA离心柱 MinElute DNA离心柱 MinElute DNA离心柱
起始材料 gDNA gDNA gDNA 全血、细胞、组织 FFPE组织切片
起始材料用量 1 ng – 2 µg gDNA 1 ng – 2 µg gDNA 1 ng – 2 µg gDNA 10–100,000个细胞,0.5–20 µl全血 1片FFPE组织切片(10 µm厚)
是否包括样本裂解  否  否 是,含有可裂解细胞、血液和组织的Lysis Buffer FTB 是,含有Deparaffinization Solution和Lysis Buffer FTB
洗脱体积 20–40 µl 40–50 µl ≥10 µl ≥10 µl ≥10 µl
转化效率 >99% >99% >99% >99% >99%
自动化 QIAcube全自动核酸纯化仪 QIAcube全自动核酸纯化仪 QIAcube全自动核酸纯化仪 QIAcube全自动核酸纯化仪
离心/真空 离心 离心/真空 离心 离心 离心
试剂盒组成 EpiTect Bisulfite Kit EpiTect 96 Bisulfite Kit EpiTect Plus DNA Bisulfite Kit EpiTect Plus LyseAll Lysis Kit和EpiTect Plus DNA Bisulfite Kit EpiTect Plus FFPE Lysis Kit和EpiTect Plus DNA Bisulfite Kit
Procedure
EpiTect Plus DNA Bisulfite Kit

EpiTect Plus DNA Bisulfite Kit含有特别配制的缓冲液和试剂,可对基因组DNA进行高效的亚硫酸氢盐转化和回收(参见"EpiTect Plus DNA flowchart")。该试剂盒具有独特的DNA保护技术和QIAGEN离心柱,可确保完全转化DNA、降解程度最低,且可洗脱到10 µl的缓冲液中,适合需要高浓度DNA的下游操作。该试剂盒可用于从20 µl溶液中转化1 ng–2 µg DNA或40 µl溶液中转化1–500 ng DNA。

EpiTect Plus FFPE Bisulfite Kit

EpiTect Plus FFPE Bisulfite Kit包括用于去除石蜡和裂解FFPE组织切片的EpiTect Plus FFPE Lysis Kit和用于对样本DNA进行高效亚硫酸氢盐转化的EpiTect Plus DNA Bisulfite Kit(参见"EpiTect Plus FFPE flowchart")。使用新型的Deparaffinization Solution和Lysis Buffer FTB,在优化的温度下,对FFPE组织切片进行去石蜡化、裂解和去交联操作。裂解和去交联后的DNA可直接用EpiTect Plus DNA Bisulfite Kit进行亚硫酸氢盐转化。该试剂盒的起始材料为厚度为10 µm的单片FFPE组织切片。

EpiTect Plus LyseAll Bisulfite Kit

EpiTect Plus LyseAll Bisulfite Kit包括用于裂解全血细胞、培养细胞和新鲜组织的EpiTect Plus LyseAll Lysis Kit和用于对样本DNA进行高效亚硫酸氢盐转化的EpiTect Plus DNA Bisulftie Kit(参见"EpiTect Plus LyseAll flowchart")。特别配制的裂解缓冲液可更高效的进行组织裂解,得到的DNA可直接用于亚硫酸氢盐转化,无需单独的纯化步骤。该试剂盒适用于0.5–20 µl血液或10–105个细胞(至少60 pg DNA)。

Applications

使用EpiTect Plus Bisulfite Kit转化和纯化的DNA适用于多种下游应用,包括:

  • 甲基化特异性PCR
  • 多重PCR
  • Real-time PCR
  • 测序/焦磷酸测序

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