title: Multiplexed optical coding nanobeads and their application in single-molecule counting analysis for multiple gene expression analysis.
abstract: A method for fabrication of multiplexed optical coding nanobeads (MOCNBs) was developed by hybridizing three types of coding DNAs labeled with different dyes (Cy5, FAM and AMCA) at precisely controlled ratios with biotinylated reporter DNA modified to magnetic streptavidin-coated nanobeads with a diameter of 300 nm. The color of the MOCNBs could be observed by overlapping three single-primary-color fluorescence images of the MOCNBs corresponding to emission of Cy5 (red), FAM (green) and AMCA (blue). The MOCNBs could be easily identified under a conventional fluorescence microscope. The MOCNBs with different colors could serve as the multiplexed optical coding labels for single-molecule counting analysis (SMCA) and be used in multi-gene expression analysis (MGEA). In the SMCA-based MGEA technique, multiple messenger RNAs (mRNAs) in cells could be simultaneously quantified through their complementary DNAs (cDNAs) by counting the bright dots with the same color corresponding to the single cDNA molecules labeled with the MOCNBs. We measured expression profiles of three genes from Lepidoptera insect Helicoverpa armigera in ∼100 HaEpi cells with and without steroid hormone inductions to demonstrate the SMCA-based MGEA technique using MOCNBs.
authors: School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China.
authors: Lu
authors: L
authors: Li
authors: School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China.
authors: Liping
authors: L
authors: Shen
authors: School of Life Science, Shandong University, Jinan 250100, China.
authors: Xiaoqian
authors: X
authors: Zhang
authors: South China Academy of Advanced Optoelectronics, South China Normal University, Guangzhou 510006, China. Electronic address: shuill@scnu.edu.cn.
authors: Lingling
authors: L
authors: Shui
authors: School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China.
authors: Benhui
authors: B
authors: Sui
authors: School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China.
authors: Xiaoli
authors: X
authors: Zhang
authors: School of Life Science, Shandong University, Jinan 250100, China. Electronic address: xfzhao@sdu.edu.cn.
authors: Xiaofan
authors: X
authors: Zhao
authors: School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China. Electronic address: jwr@sdu.edu.cn.
authors: Wenrui
authors: W
authors: Jin
mesh_terms: D000818:Animals; D019921:Biotinylation; D002460:Cell Line; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry*; D020869:Gene Expression Profiling / Q000379:methods*; D007915:Lepidoptera / Q000737:chemistry / Q000235:genetics*; D008856:Microscopy, Fluorescence; D036103:Nanotechnology; D009693:Nucleic Acid Hybridization; D045663:Quantum Dots; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics*; D059467:Transcriptome
chemical_list: D005456:Fluorescent Dyes; D012333:RNA, Messenger; D004247:DNA
keywords: Deoxyribonucleic acid determination; Multiple gene expression; Multiplexed optical coding nanobeads; Single-molecule counting analysis