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Plate-based single-cell RNA sequencing

FP Frank Penkava
MV Martin Del Castillo Velasco-Herrera
MY Matthew D. Young
NY Nicole Yager
LN Lilian N. Nwosu
AP Arthur G. Pratt
AL Alicia Lledo Lara
CG Charlotte Guzzo
AM Ash Maroof
LM Lira Mamanova
SC Suzanne Cole
ME Mirjana Efremova
DS Davide Simone
AF Andrew Filer
CB Chrysothemis C. Brown
AC Andrew L. Croxford
JI John D. Isaacs
ST Sarah Teichmann
PB Paul Bowness
SB Sam Behjati
MA M. Hussein Al-Mossawi
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Freshly sorted CD45RA-negative CD3+CD4+, and CD3+CD8+ single cells from four patients were individually flow-sorted into 96-well full-skirted plates (Eppendorf) containing 10 µL of 2% Dithiothreitol (DTT, 2 M Sigma-Aldrich), RTL lysis buffer (Qiagen) solution. Cell lysates were sealed, mixed and spun down before storing at −80 °C. Paired-end multiplexed sequencing libraries were prepared following the Smart-seq 2 (SS2) protocol33 using the Nextera XT DNA library prep kit (Illumina). A pool of barcoded libraries from four different plates were sequenced across two lanes on the Illumina HiSeq 2500.

Copyright and License information: The Author(s) ©2020
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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