The use of the restriction enzymes eliminates the problem of incomplete enzyme digestion; hnce, gving accurate findings. Te limitation of the technique includes that incomplete bisulfite modification of the DNA gives false positivity results (Szyf, 2010 p 56). Frther, por design of the primers could give inconclusive results. Bsulfite sequencing technique entails the use of bisulphite treatment of the DNA to establish its pattern of methylation. I is a qualitative technique and was the first epigenetic mark established. Te treatment of the DNA with bisulphite converts the cytosine bt does not change the 5-methylcytosine.
Te bisulphite treatment introduces specified changes to the DNA sequence basing on the methylation status of the given CpG site in the study. Te process allows for objective analysis of the nucleotide to obtain information of the methylation status of the segment of DNA in the study (Sulewska, Nklinska, Kzlowski, Mnarowski, Nklinski, Dbrowska & Chyczewski, 2007 p 319). Te Bisulfite sequencing applies a routine sequencing process on the genomic DNA treated with bisulphite allowing for the establishment of methylation status at individual CpG the strategic processes applied assume that the bisulphite induced in the genome facilitated complete conversion of the unmethylated cytosine to uracil (Hong, Cement, Cement, Hmmoud, Carrell, Cirns, Sell, Cement & Johnson, 2013 p 337).
Te key advantage of this technique established is the sensitivity it gives at each degree of methylation. I each position of the cytosine in study, i is easy to identify the degree of methylation with immense precision. Tis precision accuracy occurs since the process delineates the methylation status of each given CpG site, alowing ease identification (Szyf, 2010 p 68).
Te limitations of the technique include that it is technically demanding. Mreover, i is time-consuming and labor intensive. Tus, i is essential to consider these factors in using the method for obtaining information on DNA methylation. Te heterogeneity of cells in observing the DNA methylation status at specific CpG site becomes a problem when assessing the methylation status. Terefore, te methylation-sensitive restriction enzyme PCR is a two-step technique for the analysis of the cells in displaying methylation at given CpG in the of specific Te initial step entails. ..
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