The development of a convenient and efficient assay using miRNA-21 as a lung cancer marker is of great importance for the early prevention of cancer. Herein, an electrochemical biosensor for the detection of miRNA-21 was successfully fabricated under blue light excitation using click chemistry and photocatalytic atom transfer radical polymerization (photo-ATRP). By using hairpin DNA as a recognition probe, the electrochemical sensor deposits numerous electroactive monomers (ferrocenylmethyl methacrylate) on the electrode surface under the reaction of photocatalyst (fluorescein) and pentamethyldiethylenetriamine, thereby achieving signal amplification. This biosensor is sensitive, precise and selective for miRNA-21, and is highly specific for RNAs with different base mismatches. Under optimal conditions, the biosensor showed a linear relationship in the range of 10 fM ∼1 nM (R2 = 0.995), with a detection limit of 1.35 fM. Furthermore, the biosensor exhibits anti-interference performance when analyzing RNAs in serum samples. The biosensor is based on green chemistry and has the advantages of low cost, specificity and anti-interference ability, providing economic benefits while achieving detection objectives, which makes it highly promising for the analysis of complex samples.Copyright © 2023 Elsevier B.V. All rights reserved.