We describe the synthesis and properties of five dinucleotide fluorescent cap 
analogues labelled at the ribose of the 7-methylguanosine moiety with either 
anthraniloyl (Ant) or N-methylanthraniloyl (Mant), which have been designed for 
the preparation of fluorescent mRNAs via transcription in vitro. Two of the 
analogues bear a methylene modification in the triphosphate bridge, providing 
resistance against either the Dcp2 or DcpS decapping enzymes. All these 
compounds were prepared by ZnCl2-mediated coupling of a nucleotide P-imidazolide 
with a fluorescently labelled mononucleotide. To evaluate the utility of these 
compounds for studying interactions with cap-binding proteins and cap-related 
cellular processes, both biological and spectroscopic features of those 
compounds were determined. The results indicate acceptable quantum yields of 
fluorescence, pH independence, environmental sensitivity, and photostability. 
The cap analogues are incorporated by RNA polymerase into mRNA transcripts that 
are efficiently translated in vitro. Transcripts containing fluorescent caps but 
unmodified in the triphosphate chain are hydrolysed by Dcp2 whereas those 
containing a α-β methylene modification are resistant. Model studies exploiting 
sensitivity of Mant to changes of local environment demonstrated utility of the 
synthesized compounds for studying cap-related proteins.