Optimizing the SYBR green related cyanine dye structure to aim for brighter nucleic acid visualization

Modifying the terminal phenyl group of monomethine cyanine dyes as a pathway to brighter nucleic acid probes

“…The plane of the phenyl ring shows angles of 67.01 in 1 and of 71.61 (72.71, 2nd molecule) in 3 to the plane of the dye skeleton. The high-level DFT [M06-2X, def2-TZVP with acetonitrile (dielectric = 37.50] as a solvent using C-PCM model) calculations match extremely well with the SCXRD geometry of OxN-NMe 2 (1), OxN-OMe (3) and previously 16 studied dye PyrON (5) (Fig. S19, ESI †).…”

“…OxN (4) and OxN-OMe (3) have slightly higher exaltation (ratio of intensity) with RNA than SYBR Green II , but the intensities are quite similar. PyrON (5) and PyrN (6) presented in our previous studies, 16 have significantly lower exaltation compared to the commercial dyes SYBR Green I and II , but a slight preference could be seen – PyrON (5) has higher exaltation with DNA and PyrN (6) with RNA, respectively. The only structural difference between these two is the change from oxazolium ( PyrON (5) ) to thiazolium moiety ( PyrN (6) ).…”

“…14 newly synthesized monomethine cyanine dyes and their structural differences and their effect to the photophysical qualities have been studied by us. [14][15][16] The goal is to identify the important elements, so that the best structural features can be combined to produce the ultimate fluorescent probe for nucleic acid staining. In our previous studies, the binding properties were investigated for all 14 dyes with DNA and for selected dyes, also with RNA, to understand the phenomena behind the great turnon fluorescence.…”

“…Dimethylamine as a substituent was obvious choice since we had already seen that it improves the binding and brightness compared to thiol 15,16 and since it its capable of hydrogen bonding with the lone pair electrons, it could also improve the water solubility and at the same time enhance the binding affinity with the DNA. PEG group was included to for certain to see if the water solubility could be the answer to brighter and perhaps more user-friendly usage.…”

“…All dyes (1-4) have the highest absorption coefficients in ethanol (Table 1), following the trend previously observed. 15,16 Interestingly, OxN-NMe 2 (1) and OxN-PEG (2) have the lowest absorption coefficients in TE buffer. This could potentially indicate that without DNA in solution, dyes can for example aggregate, which is known quality of cyanine dyes, 26 lowering the absorption coefficients.…”