title: Label-free and dual-amplified detection of protein via small molecule-ligand linked DNA and a cooperative DNA machine.
abstract: Sensitive detection of protein is essential for both molecular diagnostics and biomedical research. Here, taking folate receptor as the model analyte, we developed a label-free and dual-amplified strategy via small molecular-ligand linked DNA and a cooperative DNA machine which could perform primary amplification and mediate secondary amplification simultaneously. Firstly, the specific binding of folate receptor to the small-molecule folate which linked to a trigger DNA could protect the trigger DNA from exonuclease I digestion, translating folate receptor detection into trigger DNA detection. Subsequently, trigger DNA initiated the DNA machine through hybridizing with the hairpin of the DNA machine, resulting in hairpin conformational change and stem open. The open stem further hybridized with a primer which initiated circular strand-displacement polymerization reaction; meanwhile the rolling circle amplification templates which were initially blocked in the DNA machine were liberated to mediate rolling circle amplification. In such a working model, the DNA machine achieved cooperatively controlling circular strand-displacement polymerization reaction and rolling circle amplification, realizing dual-amplification. Finally, the rolling circle amplification process synthesized a long repeated G-quadruplex sequence, which strongly interacted with N-methyl mesoporphyrin IX, bringing label-free fluorescence signal. This strategy could detect folate receptor as low as 0.23 pM. A recovery over 90% was obtained when folate receptor was detected in spiked human serum, demonstrating the feasibility of this detection strategy in biological samples.
authors.affiliation: Key Laboratory for Colloid and Interface Chemistry of Education Ministry, School of Chemistry and Chemical Engineering, Shandong University, 250100 Jinan, PR China.
authors.forename: Pei
authors.initials: P
authors.lastname: Li
authors.affiliation: School of Pharmacy, Shandong University, Jinan 250012, PR China.
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.affiliation: Key Laboratory for Colloid and Interface Chemistry of Education Ministry, School of Chemistry and Chemical Engineering, Shandong University, 250100 Jinan, PR China.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Key Laboratory for Colloid and Interface Chemistry of Education Ministry, School of Chemistry and Chemical Engineering, Shandong University, 250100 Jinan, PR China.
authors.forename: Yushu
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Key Laboratory for Colloid and Interface Chemistry of Education Ministry, School of Chemistry and Chemical Engineering, Shandong University, 250100 Jinan, PR China. Electronic address: wjiang@sdu.edu.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Jiang
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D058973:Folate Receptors, GPI-Anchored / Q000032:analysis* / Q000097:blood / Q000378:metabolism; D005492:Folic Acid / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008024:Ligands; D057230:Limit of Detection; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
chemical_list: D058973:Folate Receptors, GPI-Anchored; D008024:Ligands; D004247:DNA; D005492:Folic Acid
keywords: Cooperative DNA machine; Dual-amplification; Folate receptor; Label-free; Small molecular-ligand linked DNA.