Chaperone probes and bead-based enhancement to improve the direct detection of mRNA using silicon photonic sensor arrays.
Herein, we describe the utility of chaperone probes and a bead-based signal enhancement strategy for the analysis of full length mRNA transcripts using arrays of silicon photonic microring resonators. Changes in the local refractive index near microring sensors associated with biomolecular binding events are transduced as a shift in the resonant wavelength supported by the cavity, enabling the sensitive analysis of numerous analytes of interest. We employ the sensing platform for both the direct and bead-enhanced detection of three different mRNA transcripts, achieving a dynamic range spanning over 4 orders of magnitude and demonstrating expression profiling capabilities in total RNA extracts from the HL-60 cell line. Small, dual-use DNA chaperone molecules were developed and found to both enhance the binding kinetics of mRNA transcripts by disrupting complex secondary structure and serve as sequence-specific linkers for subsequent bead amplification. Importantly, this approach does not require amplification of the mRNA transcript, thereby allowing for simplified analyses that do not require expensive enzymatic reagents or temperature ramping capabilities associated with RT-PCR-based methods.
Department of Chemistry, University of Illinois at Urbana-Champaign, 61801, USA.
Jared T
JT
Kindt
Ryan C
RC
Bailey
D015374:Biosensing Techniques; D018922:HL-60 Cells; D006801:Humans; D007700:Kinetics; D046228:Microarray Analysis; D008863:Microspheres; D015335:Molecular Probes / Q000737:chemistry*; D053758:Nanoparticles / Q000737:chemistry; D017785:Photons; D012333:RNA, Messenger / Q000032:analysis*; D012825:Silicon / Q000737:chemistry*
D015335:Molecular Probes; D012333:RNA, Messenger; D012825:Silicon