Silver nanoclusters (AgNCs) generated on DNA templates belong to a new class of fluorescent tags showing excellent brightness, photostability and biocompatibility. Thus, AgNCs-DNA has been applied in various applications, from the detection of DNA/RNA and environmental monitoring to bioimaging and cancer therapy. In this work, we report fluorescent AgNCs synthesized using two 1,3-diaza-2-oxophenothiazine (tC)-modified oligonucleotides that contain RET-related sequence CCCCGCCCCGCCCCGCCCCA. The communication compares the absorption and emission properties of the obtained systems with silver nanoclusters synthesized on the unmodified oligonucleotide. First, we showed the optimal conditions for AgNCs-DNA synthesis on three DNA templates: (1) RET20 with the sequence 5′-CCC CGC CCC GCC CCG CCC CA-3′; (2) RET19tC with the sequence 5′-CCC CGC CCC GCC CCG CCC tCA-3′; and (3) RET14tC with the sequence 5′-CCC CGC CCC GCC CtCG CCC CA-3′. Next, the silver nanoclusters were characterized by UV/Vis absorption, fluorescence and circular dichroism spectroscopy. Silver nanoclusters RET19tC-AgNCs and RET14tC-AgNCs indicated several times higher fluorescence intensities in the long-wave emission spectra as compared to RET20-AgNCs. Moreover, silver nanoclusters on tC-modified oligonucleotides showed higher stability over time. The possibility of using the silver nanoclusters RET19tC-AgNCs for monitoring pH changes will be also tested.