The reaction kinetics and yield of traditional DNA assembly
with
a low local concentration in homogeneous solution remain challenging.
Exploring confined catalytic DNA assembly (CCDA) is intriguing to
boost the reaction rate and efficacy for creating rapid and sensitive
biosensing platforms. A rolling circle amplification (RCA) product
containing multiple tandem repeats is a natural scaffold capable of
guiding the periodic assembly of customized functional probes at precise
sites. Here, we present a RCA-confined CCDA strategy to speed up amplifiable
conversion for ratiometric fluorescent sensing of a sequence-specific
inducer (I*) by using string green-/red-Ag
clusters (sgAgCs and srAgCs) as
two counterbalance emitters. Upon recognition of I*, CCDA events are operated by two toehold-mediated strand
displacements and localized in repetitive units, thereby releasing I* for recycled signal amplification in the
as-grown RCA concatemer. The local concentration of reactive species
is increased to facilitate rapider dsDNA complex assembly and more
efficient input–output conversion, on which the clustering
template sequences of sgAgCs and srAgCs are blocked and opened, enabling srAgCs synthesis
but opposite to sgAgCs. Thus, the fluorescence emission
of srAgCs goes up, while sgAgCs
go down. With the resultant ratio featuring inherent built-in correction,
rapid, sensitive, and accurate quantification of I* at the picomolar level is achieved. Benefiting from efficient
RCA confinement to enhance reaction kinetics and conversion yield,
this CCDA-based strategy provides a new paradigm for developing simple
and diverse biosensing methodologies.