Single tube bead-based DNA co-barcoding for cost effective and accurate sequencing, haplotyping, and assembly

Single tube long fragment read (stLFR) technology enables efficient WGS, haplotyping, and contig scaffolding. It is based on adding the same barcode sequence to sub-fragments of the original DNA molecule (DNA co-barcoding). To achieve this, stLFR uses the surface of microbeads to create millions of miniaturized compartments in a single tube. Using a combinatorial process over 1.8 billion unique barcode sequences were generated on beads, enabling practically non-redundant co-barcoding in reactions with 50 million barcodes. Using stLFR we demonstrate efficient unique co-barcoding of over 8 million 20-300 kb genomic DNA fragments with near perfect variant calling and phasing of the genome of NA12878 into contigs up to N50 23.4 Mb. stLFR represents a low-cost single library solution that can enable long sequence data.