Epigenetics is the study of interaction between genes and their expressed product which is controlled by some specific chemical modification. it has a vital role in developmental process in mammalian and regulation of gene expression.dna methylation occurs most often in cpg islands (cpg sites in dna) in the promoter of genes so it plays an important role in gene expression activity. during this process, dna methyltransferases enzyme adds methyl groups to the carbon-5 position of cytosine base in the dna. abberant forms of dna methylation can cause malignancy and disorders such as different cancers [1, 2] so dna methylation detection can help early diagnosis of cancer and other related disease. many attempts have been made for dna methylation detection and among them bisulfite treatments is most common method that relies on pcr amplification [3] however recently, there have been many efforts to develop novel methods to avoid bisulfite treatment or even pcr[4-8] .all of these methods were expensive and time-consuming and there was the possibility of frequent false positives results,so the researchers efforts focused on finding the simple ,low cost and reliable methods. graphene quantum dot is a new generation of 0 dimensial carbon materials which is so popular because of unique photoluminescence properties, photostability ,biocompatibility water solubility and less toxicity ,in comparison with organic dyes and semi- conductor quantum dots (semi-qds)[9]in this work we study the interaction of grapheme quantum dot as a probe with methylated and unmethylated dna in order to show the difference between them.
Methods
In this study we prepared gqds through bottom-up approach according to yan et al [10]. all oligonucleotides stock solutions were prepared with te buffer and kept frozen until used. to make a te buffer, 1 ml of 1 m tris-hcl (ph 7.5) and 0.2 ml edta (0.5 m) was added to deionized water to total volume of 100 ml of solution. all chemicals were of analytical grade and used without further purification.
Results
the gqds were characterized by tem and the results showed that gqds were distributed in the range of 15-25 nm (fig.1). the interaction of both methylated and unmethylated dna with gqds was measured by florescence spectroscopy. the results showed that the intensity of fluorescence of the gqds at 476 nm (under excitation at 380 nm) increased after incubation with unmethylated dna. it is assumed that electron-hole recombination or zig-zag effect increases the gqds fluorescence intensity .the zigzag edge sites of quantum-sized gqds are carbene-like with a triplet ground state. hence the fluorescence intensity of gqds is efficiently enhanced. it seems that intercalation is the likely interaction mode between gqds and dna, this result also was reported previously by lu et al [11]. the fluorescence intensity dramatically decreased with the addition of methylated dsdna (fig.2) it is supposed that the presence of methyl group can change the net charge of the gqds donation of electeron which leads to a decrease in fluorescence intensity. regarding these results illustrated that this method could discriminate between methylated and unmethylated dsdna.
Conclusion
We studied the interaction of cpg rich methylated and unmethylated dna hybrids with gqds as an optical probe via fluorescence spectrometry. it was shown that methylated and unmethylated dsdna made changes in the fluorescence intensity of gqd so that under optimum conditions it could be possible to distinguish methylated from unmethylated dsdna.
Keywords
dna methylation, grapheme quantum dots, fluorescence, detection
