An improved method of bisulfite treatment and purification to study precise DNA methylation from as little as 10 pg DNA.
Methylation of vertebrate DNA is one of the most important epigenetic alterations which have become a center of scientific attraction, especially because of its important role in the regulation of transcription, genomic imprinting, developmental process, and pathogenesis of various diseases. Currently, there are wide ranges of methods available to produce quantitative and qualitative information on genomic DNA methylation. The vast majority of these methods rely on the optimization of the efficient bisulfite treatment. However, all the available methods for bisulfite treatment suffer from major disadvantages, such as large amount of starting material, poor conversion efficiency as well as low recovery and integrity of DNA after bisulfite treatment. Here, we developed a simple, rapid, and convenient column-based bisulfite treatment method by improving the several critical steps, which leads to consistent C-to-U conversion rate 99-100 %, >75 % recovery of DNA after bisulfite treatment. In addition, it is commercially viable and requires very less amount (~10 pg) of DNA.
Animal Genomics Lab, ABTC, NDRI, Karnal 132001, India.
Sandeep K
SK
Rajput
Sandeep
S
Kumar
Vivek P
VP
Dave
Ankita
A
Rajput
Haushila P
HP
Pandey
Tirtha K
TK
Datta
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification*; D019175:DNA Methylation / Q000187:drug effects*; D056426:Genetic Loci / Q000235:genetics; D005821:Genetic Techniques; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D013447:Sulfites / Q000494:pharmacology*; D013997:Time Factors
D013447:Sulfites; D004247:DNA; C042345:hydrogen sulfite