mosquito HTS: miniaturising single-cell next-generation sequencing (NGS) library preparations

Novel technologies for single-cell next-generation sequencing (NGS) offer new opportunities for understanding variations among the genomes, epigenomes and transcriptomes of seemingly identical cells. As the cost of NGS has decreased, the library preparation cost has become a larger portion of the total expenditure. This is especially true for high-throughput applications, such as single-cell analysis in areas of stem cell and cancer research. Therefore, there is a need to develop methods that can not only study the transcriptomes of single cells but can also feasibly analyse large numbers of single cells. Miniaturising the sample preparation volume provides the opportunity for significant cost savings with reductions in both reagent and sample quantities.

Nextera XT and Nextera sample prep kits (Illumina, Inc., USA) are commonly used to prepare DNA libraries. Having a precise and accurate ratio of tagmentation enzyme to DNA is critical in obtaining libraries with correct insert sizes. To assure high accuracy and precision, most library preparation protocols recommend volumes that are within the range of manual pipettes or that of large volume liquid handlers. Only a small proportion of each of the libraries prepared will be required for the sequencing, therefore reducing the library prep volume, which would also dramatically reduce the amount of sample input to pg vales, will reduce costs. This method has proven essential, particularly in high-throughput applications such as single-cell analysis, where both sample prep cost and limited starting material are major bottlenecks.

This application note presents examples of how TTP Labtech’s mosquito® HTS liquid handler can help overcome the limitations of sample prep cost associated with high-throughput applications, such as single-cell sequencing.