Nanopore PCR-free direct cDNA sequencing of human kidney tissue
Anonynous donor kidneys were sequenced using the Nanopore long-read direct cDNA sequencing kit (SQK-DCS109) and R9.4.1 flow cells. Overall design: Total RNA was extracted from frozen kidney tissues (anonymous human donors) using the Zymo Direct-zol RNA miniprep kit, including on-column DNase I digetsion. PolyA mRNA enrichment was performed using the Dynabeads mRNA DIRECT Micro kit. Reverse transcription and strand-switching were carried out using Maxima H Minus Reverse Transcriptase following the Nanopore protocol (SQK-DCS109). After end-prep, adapter ligation, and AMPure XP bead binding, sequencing was conducted on R9.4.1 flow cells using the GridIon platform.