Integration of Droplet Microfluidic Tools for Single-Cell Functional Metagenomics: An Engineering Head Start / 基因组蛋白质组与生物信息学报·英文版

Droplet microfluidic techniques have shown promising outcome to study single cells at high throughput. However, their adoption in laboratories studying "-omics" sciences is still irrelevant due to the complex and multidisciplinary nature of the field. To facilitate their use, here we provide engineering details and organized protocols for integrating three droplet-based microfluidic technologies into the metagenomic pipeline to enable functional screening of bioproducts at high throughput. First, a device encapsulating single cells in droplets at a rate of ∼250 Hz is described considering droplet size and cell growth. Then, we expand on previously reported fluorescence-activated droplet sorting systems to integrate the use of 4 independent fluorescence-exciting lasers (i.e., 405, 488, 561, and 637 nm) in a single platform to make it compatible with different fluorescence-emitting biosensors. For this sorter, both hardware and software are provided and optimized for effortlessly sorting droplets at 60 Hz. Then, a passive droplet merger is also integrated into our pipeline to enable adding new reagents to already-made droplets at a rate of 200 Hz. Finally, we provide an optimized recipe for manufacturing these chips using silicon dry-etching tools. Because of the overall integration and the technical details presented here, our approach allows biologists to quickly use microfluidic technologies and achieve both single-cell resolution and high-throughput capability (>50,000 cells/day) for mining and bioprospecting metagenomic data.

Ribosome Profiling for Biomarker Discovery / 日本補完代替医療学会誌

It has been well known that protein level is estimated by the expression level of its mRNA. However, it is also argued that correlation between mRNA abundance and protein levels is weaker than previously thought. Recently a newly developed technique called ribosome profiling has drawn attention as a drastic countermeasure to improve the weak correlation. Here it is discussed that weak association of protein and mRNA levels seen in genome-wide analysis of gene expression such as microarray is attributable to post-transcriptional regulation including translational inhibition. This review further discusses how these issues are resolved by ribosome profiling and also addresses a possibility of biomarker discovery derived from this technique.<br>