NGS Library Preparation
Low-volume dispensing and miniaturization of library preparation for next-generation sequencing.
Next-generation sequencing (NGS), a high-throughput method of single-cell sequencing, is compatible with a variety of samples, including microbial genomes and transcriptomes. Prior to sequencing, the genomes are fragmented into smaller sizes, and adapters are introduced for flow cell attachment during sequencing. These adapters can contain barcodes or indexes for multiplexing. NGS library prep of samples is an important final step before sequencing.
As NGS library prep is a significant cost of the total workflow, miniaturization of library prep enables the generation of high-quality samples while reducing overall costs. Cost also becomes more critical when many libraries are sequenced simultaneously. The I.DOT provides easy reaction miniaturization, while maintaining flexibility to perform various types of NGS library protocols.
How the I.DOT facilitates NGS library prep
Low volume non-contact dispensing.
Dispendix’s portfolio features the Immediate Drop-on-demand Technology (I.DOT), a patented approach for optimizing non-contact liquid-handling tasks. Specifically, the I.DOT provides easy reaction miniaturization, while maintaining flexibility to perform various types of NGS library protocols. The system uses eight individually controlled positive pressure channels to generate droplets from 8 to 50 nL from a small outlet at the bottom of each well. Each channel can generate up to 100 droplets per second, which enables a volume range from 8 nL to 80 μL. The non-contact dispensing significantly reduces the number of pipette tips required during NGS library prep, and the low-volume dispensing (dead volume <1 µL) provides significant cost savings on reagents.
Quantification and Normalization of NGS libraries
Typically, complementary DNA (cDNA) amplification from single cells results in uneven concentrations, which may require normalization to ensure an even read distribution for all cells. Hundreds of libraries can be quantified and normalized within minutes using the I.DOT, which allows for highly multiplexed dispensing of up to 96 reagents in parallel.
LIBRARY INDEXING AND EARLY BARCODING
The ability to index NGS libraries with unique DNA barcodes provides the basis for high-throughput single-cell sequencing. For single-cell full-length transcript sequencing or whole exome sequencing, individual libraries are usually indexed after amplification and fragmentation.
PERFORMING COMPLEX TASKS AT UNRIVALED SPEED
The I.DOT non-contact dispensing technology was developed to perform even the most complex liquid-handling tasks at unrivaled speeds. Up to 2304 libraries can be uniquely barcoded with a single I.DOT source plate. Dispensing 96 unique barcodes takes less than 60 seconds.