16S rRNA gene amplicon (metabarcoding) of surface-sterilized root nodules from Pisum sativum (yellow pea)
Pisum sativum (yellow pea) root nodules were collected at anthesis, surface-sterilized (70% ethanol 1 min; 2% sodium hypochlorite 23 min; three sterile water rinses), and macerated. Total DNA was extracted (QIAGEN DNeasy Plant Pro Kit). Full-length 16S rRNA libraries were prepared with the Oxford Nanopore 16S Barcoding Kit (SQK-16S024) per manufacturers instructions, including PCR with kit primers, barcode attachment, cleanup (AMPure XP), quantification (Qubit), and equimolar pooling. Libraries were sequenced on an Oxford Nanopore. Raw signals were basecalled with ONT Dorado; demultiplexing was performed with `dorado demux`.