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Acknowledgements
We thank A. Morgan for technical assistance. This study was supported by grants DP1EY023176, P30EY002520, T32EY07001,and DP1OD008301 to A.S.T.; grants from the Swedish Research Council and the Swedish Foundation for Strategic Research (FFL4) to R.S.; grant R01MH103108 to A.S.T. and K.F.T.; grant R01NS062829 to K.F.T.; the McKnight Scholar Award to A.S.T.; and the Arnold and Mabel Beckman Foundation Young Investigator Award to A.S.T. C.R.C. was supported by grants F30MH095440, T32GM007330 and T32EB006350. M.B. and P.B. were supported by the Deutsche Forschungsgemeinschaft (DFG, EXC 307) and the German Federal Ministry of Education and Research (BMBF; BCCN Tübingen, FKZ 01GQ1002).
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Competing interests
R.S. has developed and patented Smart-seq2 and licensed that technology to Clontech, a Takara Bio Company.
Integrated supplementary information
Supplementary Figure 1 Optimization of Patch-seq protocol.
Pilot experiments were carried out to test the effect of various protocol modifications on cDNA yield, including addition of RNase inhibitor to the intracellular solution (a), silanization of glass capillaries (b), concentration of dNTPs in lysis buffer (c), extraction of cytoplasm only v. cytoplasm and nucleus (d), pipette tip size (e) (tip size inversely proportional to resistance, line represents linear regression), and volume of intracellular solution in patch pipette (f) (20 μl total reaction volume, RT-PCR performed on whole brain cDNA template).
Supplementary Figure 2 Modified RNase-free intracellular solution does not affect health of patch-clamp-recorded neurons.
Membrane potential of neurons patched with RNase-free modified intracellular solution for Patch-seq recordings. Data represent mean ± SE (n=3 neurons).
Supplementary Figure 3 Quality control of cDNA libraries.
(a) Average size and concentration (from 300-9,000 bp) of all collected samples. Only samples with more than 200 pg/μl and an average size greater than 1,500 bp were sequenced (criteria denoted by dashed lines). Libraries generated from patched cells generally contained more (b) and higher quality, full-length (c) cDNA compared to negative