EZ DNA Methylation-Gold Kit
D5006 / D5005
- Complete bisulfite conversion of DNA in less than 3 hours.
- Desulphonation and recovery of bisulfite-treated DNA with a spin column.
- Recovered DNA is ideal for downstream analyses such as PCR, endonuclease digestion, sequencing, microarrays, etc.
|Applications||Purified, converted DNA is of high-quality and well-suited for downstream processes, including library preparation for Next-generation sequencing, PCR amplification, etc.|
|Elution Volume||≥ 10 µl|
|Equipment||Microcentrifuge and thermocycler with heated lid|
|Input||500 pg - 2 µg of DNA.|
|Processing Time||3 hours|
|Sample Source||Purified genomic DNA, endonuclease-digested DNA, linearized plasmid DNA, etc. DNA should be high-quality and RNA-free.|
Q1: Tips for bisulfite primer design?
Q2: Is an incubation with desulphonation buffer for longer than 20 minutes recommended?
Q3: Does bisulfite conversion only occur in a CpG context?
Q4: Which polymerase is recommended for amplification from bisulfite converted DNA?
Q5: What is the minimum DNA size that can be recovered?
Q6: How to quantify / visualize converted DNA?
Q7: What leads to poor conversion efficiency/ low yields?
Q8: How long is bisulfite converted DNA stable at -20 °C?
Researchers from the National Institute of Plant Genome Research in New Delhi used the EZ DNA Methylation-Gold Kit to identify the level of cytosine methylation in genomic DNA from Tomato leaf curl New Delhi virus. Through DNA methylation-specific RNA silencing, the researchers aimed to target the viral genomic regions of this virus to potentially determine an alternate defense mechanism for generating transgenic plants to prevent yield loss.Sahu P et al. Post-transcriptional and Epigenetic Arms of RNA Silencing: A Defense Machinery of Naturally Tolerant Tomato Plant Against Tomato Leaf Curl New Delhi Virus. Plant Mol Biol Rep (2014) 32:1015-1029
The EZ DNA Methylation-Gold Kit was used to bisulfite convert DNA extracted from Thy-1 (+) and Thy-1 (-) lung fibroblasts and tissue. Data from methylation-specific PCR and bisulfite sequencing enabled researchers to show that epigenetic regulation of Thy-1 could be a novel and potentially reversible mechanism in fibrosis that may offer the possibility of new therapeutic options.Sanders Y et al. Thy-1 Promoter Hypermethylation A Novel Epigenetic Pathogenic Mechanism in Pulmonary Fibrosis. Am J Respir Cell Mol Biol. 2008 Nov; 39(5):610-8
Researchers from China analyzed the role of DNA methylation in the life cycle of a parasitic nematode, Trichinella spiralis and characterized the methylome of three life-cycles states in this organism. Genomic DNA of T. spiralis muscle larvae, adults, and new-born larvae was isolated and methylated cytosine levels were determined by MethylC-seq.Gao F et al. (2012) Differential DNA methylation in discrete developmental stages of the parasitic nematode Trichinella spiralis. Genome Biol. 13(10):R100.
“It makes gene specific DNA methylation analysis very simple and anyone with modest technical background can easily perform the experiment using this kit.”
- Murali B. (Centre for DNA Fingerprinting and Diagnostics)
“This protocol allows for bisulfite conversion in about 3 hours. This fast conversion is particularly relevant to our experiments, which allows us to PCR amplify and clone the products in 1 day, saving time without loss of quality.”