10x Genomics sequencing: 61L2
(1) DNA Preparation: Input DNA >=1 ng (recommended >=50 ng) with fragment length >=50 kb. Short fragments are removed via pulsed-field electrophoresis or bead-based selection to retain high-molecular-weight DNA. (2) GEM Formation (Gel Bead-in-Emulsions): Long DNA fragments, barcoded Gel Beads, and enzymes are partitioned into oil droplets using microfluidic chips to form GEMs. Each GEM encapsulates a single long DNA fragment, which is randomly fragmented (~350 bp) and tagged with a unique barcode. (3) Library Construction: Short fragments undergo end repair, A-tailing, and ligation of Illumina adapters (P5/P7). And PCR amplifies fragments while incorporating barcodes to generate libraries with unique molecular identifiers (UMIs). (4) Sequencing: Performed on Illumina platforms (PE150), with Read1 capturing barcodes and Read2 acquiring genomic sequences.