Single-cell gene variation analysis method for single gland.

Abstract

Single-cell analysis of heterogeneity has become the cutting-edge technology for profound understandings of relationships between cell populations. At present, common methods used in single cellular genomic research are mainly microfluidic technologies (Fluidigm) or based on microwells, both requiring a uniform size of cells at the entrance. However, the size of cells in specific tissues can vary from type to type. To address this issue, we need to establish a method to identify genomic features of individual cells of different sizes. In this paper, we developed a robust method in the analysis of single cellular genomic mutations among gastric tissues. Briefly, the single gastric gland was isolated from the whole tissue, and further enzymatically digested into single cells of various sizes by trypsin. These single cells were then spread on the polyethylene naphthalene slides and selected by the laser microdissection method. Whole genome amplification (WGA) and capillary electrophoresis were performed subsequently to detect single cell microsatellite. This method enabled us to detect the existence of microsatellite instability (MSI) of each single cell within the intestinal metaplasia, and to carry out a flexible and fine analysis of single cells with different sizes in tissues and glands. This reliable and practical method is well performed in both low and high-throughput genome analysis when combined with cell labeling methods, thus providing a novel and highly flexible way to study tissue heterogeneity on the single cell scale.