For CEL-Seq2 single-cell library preparation (58 (link)), a nanoliter pipetting robot (mosquito HTS, TTP Labtech) was used to reduce the CEL-Seq2 protocol original volumes by fivefold. The 384-well plates (Corning, PCR-384-RGD-C) were prepared with 240 nl of lysis buffer in every well in these final ratios: 20 nl of 10 mM deoxynucleotide triphosphate (1:12), 40 nl of 1:100,000 ERCC spike mix 1 or 2 (2:12), 140 nl of water with 0.35% Triton X-100 (7:12), and 40 nl of 25 ng/μl of uniquely barcoded polydT primers with unique molecular identifier (UMI; 2:12). Hydrophobic encapsulation barrier (1.2 μl; Vapor-Lock, QIAGEN, 981611) was lastly added to every well.
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CEL-Seq2 Single-Cell Library Prep in Nanoliter Volumes
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Corresponding Organization :
Other organizations : Max Planck Institute of Immunobiology and Epigenetics, University of Freiburg
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- Reduced CEL-Seq2 protocol original volumes by fivefold using a nanoliter pipetting robot (mosquito HTS, TTP Labtech)
- Not explicitly mentioned
- 384-well plates (Corning, PCR-384-RGD-C)
- Lysis buffer (240 nl per well)
- Deoxynucleotide triphosphate (20 nl of 10 mM per well; 1:12 ratio)
- ERCC spike mix 1 or 2 (40 nl of 1:100,000 per well; 2:12 ratio)
- Water with 0.35% Triton X-100 (140 nl per well; 7:12 ratio)
- Uniquely barcoded polydT primers with unique molecular identifier (UMI; 40 nl of 25 ng/μl per well; 2:12 ratio)
- Hydrophobic encapsulation barrier (1.2 μl; Vapor-Lock, QIAGEN, 981611) added to every well
- Positive control: ERCC spike mix 1 or 2
- Negative control: Not specified
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