RNA-seq Based Transcriptomic Analysis of Single Cyanobacterial Cells
Zixi Chen, Jiangxin Wang, Lei Chen and Weiwen Zhang
from: Cyanobacteria: Omics and Manipulation (Edited by: Dmitry A. Los). Caister Academic Press, U.K. (2017) Pages: 75-92.
Gene-expression heterogeneity among individual cells will eventually determine the fate of a bacterial population. We describe the first bacterial single-cell RNA sequencing (RNA-seq), BaSiC RNA-seq, a method integrating RNA isolation, cDNA synthesis and amplification, and RNA-seq analysis of the whole transcriptome of single cells of the cyanobacterium Synechocystis sp. PCC 6803, which typically contain approximately 5-7 femtogram of total RNA per cell. We applied the method to 3 Synechocystis single cells at 24 h and 3 single cells at 72 h after a nitrogen-starvation stress treatment, as well as their bulk-cell controls for the same conditions, to determine the heterogeneity upon the environmental stress. With 82-98% and 31-48% of all putative Synechocystis genes identified in single cells of 24 and 72 h, respectively, the results demonstrated the method could achieve good identification of the transcripts in single bacterial cells. In addition, the preliminary results from nitrogen-starved cells also showed a possible increasing gene-expression heterogeneity from 24 h to 72 h after nitrogen starvation. Moreover, preliminary analysis of single-cell transcriptomic datasets revealed that genes from the 'Mobile elements' functional category have the most significant increase of gene-expression heterogeneity under stress, which was further confirmed by single-cell RT-qPCR analysis of gene expression in 24 randomly selected cells read more ...