Microfluidic single-cell mRNA isolation and analysis.
Single-cell gene expression analysis holds great promise for studying diverse biological systems, but methodology to process these precious samples in a reproducible, quantitative, and parallel fashion remains challenging. Here, we utilize microfluidics to isolate picogram and subpicogram mRNA templates, as well as to synthesize cDNA from these templates. We demonstrate single-cell mRNA isolation and cDNA synthesis, provide quantitative calibrations for each step in the process, and measure gene expression in individual cells. The techniques presented here form the foundation for highly parallel single-cell gene expression studies.
Marcus
Joshua S
JS
Biochemistry and Molecular Biophysics, California Institute of Technology, MS 128-95, Pasadena, California 91125, USA.
Anderson
W French
WF
Quake
Stephen R
SR
D000818:Animals; D018076:DNA, Complementary / Q000138:chemical synthesis / Q000737:chemistry; D051379:Mice; D044085:Microfluidics / Q000295:instrumentation* / Q000379:methods*; D041681:NIH 3T3 Cells; D012333:RNA, Messenger / Q000032:analysis* / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity
D018076:DNA, Complementary; D012333:RNA, Messenger