Division of Evolutionary Biology, Faculty of Biology, LMU Munich, Grosshaderner Str. 2, Planegg-Martinsried 82152, Germany.
Division of Evolutionary Biology, Faculty of Biology, LMU Munich, Grosshaderner Str. 2, Planegg-Martinsried 82152, Germany; Gene Center and Department of Biochemistry, Ludwig-Maximilians-Universität München, München, Germany.
J Insect Physiol. 2023 Jun;147:104516. doi: 10.1016/j.jinsphys.2023.104516. Epub 2023 Apr 8.
Ectotherms such as insects are animals whose body temperature largely depends on ambient temperature and temperature variations provide a selection pressure affecting the geographical distribution of these species. However, over the course of evolution, some insect species managed to colonize environments characterized by various temperature ranges. Therefore, insects provide an excellent study system to investigate the basis of adaptation to temperature changes and extremes. We are generally using the vinegar fly Drosophila ananassae as a model system to investigate the genetic basis of cold tolerance. This species has expanded from its tropical ancestral range to more temperate regions resulting in a cosmopolitan, domestic distribution. Previously, we identified candidate genes significantly associated with cold tolerance in this species. We now established molecular genetic tools to assess the function of these genes. Using CRISPR/Cas9 methodology for genome editing and the PiggyBac system, the Cas9 enzyme was successfully integrated into the genome of three fly strains with different levels of cold tolerance. We further report on preliminary findings that the Cas9 integration itself did not have a consistent effect on tolerance to cold. In conclusion, we offer with our study the molecular tools that allow studying stress-related candidate genes in D. ananassae in the future. In addition, we point out and provide guidance on the challenges that come with genome editing in a non-model species.
变温动物，如昆虫，是其体温在很大程度上依赖于环境温度的动物，温度变化提供了一种选择压力，影响这些物种的地理分布。然而，在进化过程中，一些昆虫物种成功地在具有不同温度范围的环境中殖民。因此，昆虫为研究适应温度变化和极端温度的基础提供了一个极好的研究系统。我们通常使用醋蝇黑腹果蝇作为模型系统来研究耐寒性的遗传基础。该物种已经从其热带祖先范围扩展到更温和的地区，从而形成了世界性的、家庭式的分布。此前，我们在该物种中鉴定出与耐寒性显著相关的候选基因。我们现在建立了分子遗传工具来评估这些基因的功能。使用 CRISPR/Cas9 基因组编辑方法和 PiggyBac 系统，Cas9 酶成功地整合到三种具有不同耐寒性水平的果蝇品系的基因组中。我们进一步报告了初步发现，Cas9 整合本身对耐寒性没有一致的影响。总之，我们提供了研究 D. ananassae 中与应激相关的候选基因的分子工具。此外，我们指出并提供了在非模式物种中进行基因组编辑所面临的挑战的指导。
