Evaluation of potassium stress responses and identification of novel RT-qPCR reference genes in the haloarchaeon, Haloarcula marismortui .

Growth characteristics and stress responses in the halophilic archaeon, Haloarcula marismortui, have been poorly investigated and knowledge of the effects of extracellular potassium concentration on halophilic growth is limited. We report the evaluation of cellular generation times across a range of extracellular potassium concentrations to assess the organisms responses to extreme potassium stress. Our results show Haloarcula marismortui exhibits an optimal generation time of 4.19 ± 0.14 hours at an extracellular KC1 concentration of 100mM. This corresponds to an intracellular K⁺ concentration of 2.02M as determined through the use of Induction-Coupled Plasma Mass Spectrometry. Additionally, the validation of several candidate reference genes for use with RT-qPCR studies is reported. Five reference genes (16S rRNA, rpoB, pykA, polA, and rpoA) have been confirmed as being stably expressed in accordance with the Minimal Information for the Publication of Quantitative PCR Experiments (MIQE Guidelines) across several unique halophilic growth conditions.