title: A dumbell probe-mediated rolling circle amplification strategy for highly sensitive transcription factor detection.
abstract: Highly sensitive detection of transcription factors (TF) is essential to proteome and genomics research as well as clinical diagnosis. We describe herein a novel fluorescent-amplified strategy for ultrasensitive, quantitative, and inexpensive detection of TF. The strategy consists of a hairpin DNA probe containing a TF binding sequence for target TF, a dumbbell-shaped probe, a primer DNA probe designed partly complementary to hairpin DNA probe, and a dumbbell probe. In the presence of target TF, the binding of the TF with hairpin DNA probe will prohibit the hybridization of the primer DNA probe with the "stem" and "loop" region of the hairpin DNA probe, then the unhybridized region of the primer DNA will hybridize with dumbbell probe, subsequently promote the ligation reaction and the rolling circle amplification (RCA), finally, the RCA products are quantified via the fluorescent intensity of SYBR Green I (SG). Using TATA-binding protein (TBP) as a model transcription factor, the proposed assay system can specifically detect TBP with a detection limit as low as 40.7 fM, and with a linear range from 100 fM to 1 nM. Moreover, this assay related DNA probe does not involve any modification and the whole assay proceeds in one tube, which makes the assay simple and low cost. It is expected to become a powerful tool for bioanalysis and clinic diagnostic application.
authors.affiliation: Key Laboratory of Theoretical Organic Chemistry and Function Molecule, Ministry of Education, School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201, PR China.
authors.forename: Chunxiang
authors.initials: C
authors.lastname: Li
authors.affiliation: Key Laboratory of Theoretical Organic Chemistry and Function Molecule, Ministry of Education, School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201, PR China.
authors.forename: Xiyang
authors.initials: X
authors.lastname: Qiu
authors.affiliation: School of Chemistry and Chemical Engineering, Hunan Institute of Science and Technology, Yueyang 414006, PR China. Electronic address: zhqh96@163.com.
authors.forename: Zhaohui
authors.initials: Z
authors.lastname: Hou
authors.affiliation: Key Laboratory of Theoretical Organic Chemistry and Function Molecule, Ministry of Education, School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201, PR China. Electronic address: keqindeng@hnust.edu.cn.
authors.forename: Keqin
authors.initials: K
authors.lastname: Deng
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D015342:DNA Probes / Q000032:analysis* / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*; D014157:Transcription Factors / Q000032:analysis* / Q000235:genetics*
chemical_list: D015342:DNA Probes; D014157:Transcription Factors
keywords: Rolling circle amplification; TATA-binding protein; Transcription factor detection