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EZ-96 DNA Methylation-Direct MagPrep

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Cat #	Name	Size	Price	Quantity
D5045	EZ-96 DNA Methylation-Direct MagPrep	8 x 96 Rxns.	$1,391.70	- +
D5044	EZ-96 DNA Methylation-Direct MagPrep	4 x 96 Rxns.	$860.30	- +

Highlights

High throughput, complete bisulfite conversion of DNA directly from blood, tissue, cells, FFPE and LCM-derived samples
Compatible with small sample inputs – as few as 10 cells or 50 pg of DNA.
High throughput (96-well), automated desulphonation and recovery of bisulfite-treated DNA.

Documents

Supplemental Info

Product Description

The EZ-96 DNA Methylation-Direct MagPrep is a magnetic bead-based bisulfite conversion kit that allows simple and reliable DNA bisulfite conversion directly from blood, tissue, and cells without the prerequisite for DNA purification. The increased sensitivity of this magnetic bead-based bisulfite conversion kit makes it possible to amplify bisulfite-converted DNA from as few as 10 cells or 50 pg DNA. These innovations have been coupled to a magnetic bead-based clean-up for high-throughput methylation analysis. Desulphonation and clean-up of the converted DNA is performed while bound to the MagBinding Beads. This magnetic bead based bisulfite conversion kit has been designed to minimize template degradation, loss of DNA during treatment and clean-up, and to provide complete conversion of unmethylated cytosines. Recovered DNA is ideal for downstream analyses, including PCR amplification, endonuclease digestion, sequencing, microarrays, etc.

For automation scripts and support, email automation@zymoresearch.com

Documents

Technical Specifications

Applications	Purified, converted DNA is of high-quality and well-suited for downstream processes, including library preparation for Next-Generation sequencing, PCR amplification, etc.
Conversion	> 99.5%
Elution Volume	≥ 25 µl
Equipment	Thermocycler with heated lid, heating element for 96-well plate, magnetic stand.
Input	Samples containing between 50 pg to 2 µg of DNA. For optimal results, the amount of input DNA should be from 200 to 500 ng. The number of cells per standard treatment can range from 10-10⁵ cells.
Processing Time	4 hours
Recovery	> 80%
Sample Source	Blood, tissue, cells, FFPE, LCM-derived samples, purified genomic DNA, endonuclease-digested DNA, linearized plasmid DNA, etc.
Supplemental Info	Bisulfite Beginner Guide Controls for DNA Methylation Assays Bisulfite Conversion for Illumina Methylation Assays Bisulfite Primer Design Tool Link

Resources

Q1: Tips for bisulfite primer design?

https://www.zymoresearch.com/pages/bisulfite-beginner-guide
https://www.zymoresearch.com/pages/bisulfite-primer-seeker

Q2: Is incubation with Desulphonation Buffer for longer than 20 minutes recommended?

Leaving the desulphonation buffer on the column longer than recommended will cause more degradation and result in lower yields.

Q3: Does bisulfite conversion only occur in a CpG context?

Bisulfite conversion will work regardless of context, so the kits are compatible with genomic DNA derived from plants and other species with high non-CpG methylation levels.

Q4: Which polymerase is recommended for amplification from bisulfite converted DNA?

ZymoTaq DNA Polymerase has been specifically designed for use in bisulfite amplification reactions. ZymoTaq is available as a stand-alone polymerase (E2001/E2002), PreMix (E2003/E2004), and qPCR PreMix (E2054/E2055).

Q5: What is the minimum DNA size that can be recovered?

> 50 bp.

Q6: How to quantify converted DNA?

For best results, keep the method of quantification consistent before and after bisulfite treatment:

If quantifying with a NanoDrop, use dsDNA settings (50 μg/ml for Ab260 = 1.0) before treatment and use RNA settings (40 μg/ml for Ab260 = 1.0) after treatment.
If quantifying with Qubit, use a dsDNA assay before treatment and use a ssDNA assay after treatment.

Following bisulfite treatment of DNA, nonmethylated cytosine residues are converted into uracil. The recovered DNA is typically A, U, and T-rich. The original base-pairing no longer exists. Instead, the converted DNA is single stranded with limited non-specific base-pairing at room temperature. Therefore, traditional dsDNA methods of quantification will not accurately represent the amount of recovered bisulfite converted DNA.

Q7: How to visualize converted DNA?

Following bisulfite treatment, DNA will be single stranded with limited non-specific base pairing at room temperature. To visualize, run the converted DNA on an agarose gel then chill the gel on ice or in an ice bath for 30 minutes. This will force enough base-pairing to allow intercalation of the ethidium bromide for the DNA to be visible. If using a Bioanalyzer or TapeStation instrument, use RNA kits and reagents to visualize the converted DNA.

Q8: What leads to poor conversion efficiency/ low yields?

Poor conversion efficiency and low yields can be due to a variety of different experiment-specific conditions. Please contact Technical Support to discuss your specific experimental conditions and further troubleshoot with a product specialist.

Q9: How long is bisulfite converted DNA stable at -20°C?

Converted DNA eluted in M-Elution Buffer can generally be stored at -20°C for 1-3 months. If longer term storage is necessary, we recommend storing at or below -70°C if possible. Bisulfite converted DNA is less stable than dsDNA; for best results, minimize freeze-thawing of converted DNA and use as soon as possible for downstream analysis.

Cat #	Name	Size	Price
D4100-5-8	MagBinding Beads	8 mL	$101.20
D4100-5-16	MagBinding Beads	16 mL	$182.60
C2003	Elution Plate	2 Plates	$21.50
C2002	Collection Plate	2 Plates	$24.90
C2005	Conversion Plate w/ Cover Foil	2 Plates/Foils	$10.30
D5021-7	M-Solubilization Buffer	18 ml	$34.60
D5021-8	M-Reaction Buffer	4ml	$26.00
D5021-9	M-Digestion Buffer (2X)	15 ml	$18.50
D5040-3	M-Binding Buffer	250 ml	$129.80
D5040-5	M-Desulphonation Buffer	80 ml	$91.50
D5040-4	M-Wash Buffer	72 ml	$84.00
D5007-6	M-Elution Buffer	8 ml	$12.40
D5006-2	M-Dilution Buffer-Gold	7 ml	$12.40
D5003-1	CT Conversion Reagent (96 Conversions)	1 Bottle	$77.90