Meng Lingning, Cao Xiaoli, Li Chuchu, Li Jia, Xie Hui, Shi Jiping, Han Mei, Shen Han, Liu Chang
Department of Laboratory Medicine, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Nanjing, Jiangsu, China.
Department of Acute Infectious Disease Control and Prevention, Jiangsu Provincial Center for Disease Control and Prevention, Nanjing, China.
Front Microbiol. 2023 Mar 13;14:1140515. doi: 10.3389/fmicb.2023.1140515. eCollection 2023.
is an opportunistic human pathogen notorious for its remarkable capacity of multi-drug resistance, and has become one of the most important model bacteria in clinical bacteriology research. Quantitative real-time PCR is a reliable method widely used in gene expression analysis, for which the selection of a set of appropriate housekeeping genes is a key prerequisite for the accuracy of the results. However, it is easy to overlook that the expression level of housekeeping gene may vary in different conditions, especially in the condition of molecular microbiology assays, where tested strains are generally cultured under the pre-set antibiotic selection pressures, and how this affects the stability of commonly used housekeeping genes remains unclear. In this study, the expression stability of ten classic housekeeping genes ( and ) under the pressure of eight laboratory commonly used antibiotics (kanamycin, gentamycin, tetracycline, chloramphenicol, hygromycin B, apramycin, tellurite, and zeocin) were tested. Results showed that the stability of housekeeping gene expression was indeed affected by the types of antibiotics added, and of course the best reference gene set varied for different antibiotics. This study provides a comprehensive summary of the effects of laboratory antibiotics on the stability of housekeeping genes in , highlighting the necessity to select housekeeping genes according to the type of antibiotics used in the initial stage of experiment.
是一种机会性人类病原体，以其显著的多重耐药能力而臭名昭著，已成为临床细菌学研究中最重要的模式细菌之一。定量实时聚合酶链反应是一种广泛用于基因表达分析的可靠方法，对于该方法而言，选择一组合适的管家基因是结果准确性的关键前提。然而，很容易被忽视的是，管家基因的表达水平在不同条件下可能会有所不同，尤其是在分子微生物学检测的情况下，受试菌株通常在预设的抗生素选择压力下培养，而这如何影响常用管家基因的稳定性仍不清楚。在本研究中，测试了十种经典管家基因（和）在八种实验室常用抗生素（卡那霉素、庆大霉素、四环素、氯霉素、潮霉素B、阿泊拉霉素、亚碲酸盐和博来霉素）压力下的表达稳定性。结果表明，管家基因表达的稳定性确实受到添加抗生素类型的影响，当然，不同抗生素的最佳参考基因集也有所不同。本研究全面总结了实验室抗生素对中管家基因稳定性的影响，强调了在实验初始阶段根据所用抗生素类型选择管家基因的必要性。
