EMD Millipore™ Chemicon™ CpGenome™ Universal DNA Modification Kit

DNA methylation profiling Gene specific profiling - HepG2 FHIT

Experiment
DNA methylation profiling Gene specific profiling - HepG2 FHIT
Product
EMD Millipore™ Chemicon™ CpGenome™ Universal DNA Modification Kit from Fisher Scientific
Manufacturer
Fisher Scientific

Protocol tips

Protocol tips
For best results, the CT Conversion Reagent should be prepared freshly and used immediately following preparation.

Publication protocol

Methylation-specific polymerase chain reaction (PCR)
Total cell DNA was isolated from the cultured cells using a QIAamp DNA Mini kit (Qiagen N.V., Venlo, The Netherlands). The genomic DNA was modified as described previously () using a DNA bisulfite modification kit (Chemicon, Temecula, CA, USA). The methylated CpG dinucleotides within a CpG island in the promoter region of FHIT were detected using the following specific PCR primer sets: methylated FHIT forward, 5-TTGGGGCGCGGGTTTGGGTTTTTACGC-3 and reverse, 5-CGTAAACGACGCCGACCCCACTA-3; and unmethylated FHIT forward, 5-TTGGGGTGTGGGTTT GGGTTTTTATG-3 and reverse, 5-CATAAACAACACCAACCCCACTA-3 (). Each PCR mixture system contained 10X PCR buffer (Qiagen N.V.), deoxynucleotide triphosphates (1.25 mmol/l), primers (0.6 mmol/l), 1 unit HotStarTaq (Qiagen N.V.), and bisulfite-modified DNA (100 ng). PCR was performed in a programmable thermal controller (MJ Research, Inc., Quebec, Canada) under the following conditions: Preheating at 94°C for 3 min, 94°C for 30 sec, 62°C for 30 sec and 72°C for 30 sec for 38 cycles and a final extension at 72°C for 7 min. The PCR products were analyzed on a 2% agarose gel.

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Discussion

Discussion

5 years ago

Author: Jody Hancock Canada

How can I preserve leukocytes for DNA methylation profiling?

I would like to preserve leukocytes for future epigenetic analysis. How can I preserve them effectively in order to perform DNA methylation profiling at a later time?

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing DNA methylation profiling Gene specific profiling - HepG2 FHIT using EMD Millipore™ Chemicon™ CpGenome™ Universal DNA Modification Kit from Fisher Scientific.

Paper title
FHIT overexpression in HepG2 hepatoma cells affects growth and cyclin D1 expression
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Manufacturer protocol

Download the product protocol from Fisher Scientific for EMD Millipore™ Chemicon™ CpGenome™ Universal DNA Modification Kit below.

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