Set a to d

Sequencing libraries were prepared using the standard Illumina Nextera XT. DNA Sample Preparation kit (Illumina, #FC-131-1096) and the combination of 384 Combinatorial Dual Indexes (Illumina-Set A to D, #FC-131-2001 to FC-131-2004). Using the Mosquito liquid handling robot, the Nextera XT chemistry was miniaturized reducing 10-times the manufacturers volume as previously described [118, 119] (Supp. Table 7). For the library preparation 500nL of the undiluted cDNA was transferred in a new 384 well-plate containing 1500nL of Tagmentation Mix (TD and ATM reagents). Accordingly, all Nextera XT reagents (NT, NPM and i5/i7 indexes) were added stepwise to a final library volume of 5µL per well. We prepared a 384 well-plate with the combination of the i5/i7 index adapters, which was used in our miniaturization protocols. The final PCR amplification was 12 cycles. After library preparation, 500nL from each well was pooled together to a final volume of ~192µl to perform a final AMPure XP bead (Beckman Coulter, #A63882) clean-up. For our single nuclei libraries, two consecutive clean-ups with a ratio of sample to bead 0,9X led to library sizes between 200 and 1000bp, with no trace of adaptors. The final libraries were assessed using a HS DNA kit in the Agilent Bioanalyzer.