Genome sequencing of bacteria after catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) and multiple displacement amplification (MDA)

We evaluated the compatibility of a widely used phylogenetic staining protocol with whole genome amplification and genome sequencing. Cultures of the freshwater Betaproteobacterium Limnohabitans sp. strain RIM47 were subjected to ethanol fixation, catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH), flow cytometric sorting and multiple displacement amplification (MDA). Sequencing libraries were generated from up to 14 replicate MDA products and analyzed in the context of controls produced from unstained cells.

本研究评估了一套通用系统发育染色方案（phylogenetic staining protocol）与全基因组扩增及基因组测序的兼容性。以淡水β-变形菌门（Betaproteobacterium）Limnohabitans属菌株RIM47的培养物为实验材料，依次经乙醇固定、催化报道分子沉积荧光原位杂交（CARD-FISH）、流式细胞分选及多重置换扩增（MDA）处理。从最多14份重复MDA扩增产物中构建测序文库，并以未染色细胞制备的对照样本为参照开展分析。