Peuß Robert, Zakibe Zachary, Krishnan Jaya, Merryman M Shane, Baumann Diana P, Rohner Nicolas
Stowers Institute for Medical Research.
Stowers Institute for Medical Research; Department of Molecular & Integrative Physiology, KU Medical Center;
J Vis Exp. 2019 May 25(147). doi: 10.3791/59334.
Astyanax mexicanus is emerging as a model organism for a variety of research fields in biological science. Part of the recent success of this teleost fish species is that it possesses interfertile cave and river-dwelling populations. This enables the genetic mapping of heritable traits that were fixed during adaptation to the different environments of these populations. While this species can be maintained and bred in the lab, it is challenging to both obtain embryos during the daytime and create hybrid embryos between strains. In vitro fertilization (IVF) has been used with a variety of different model organisms to successfully and repeatedly breed animals in the lab. In this protocol, we show how, by acclimatizing A. mexicanus to different light cycles coupled with changes in water temperature, we can shift breeding cycles to a chosen time of the day. Subsequently, we show how to identify suitable parental fish, collect healthy gametes from males and females, and produce viable offspring using IVF. This enables related procedures such as the injection of genetic constructs or developmental analysis to occur during normal working hours. Furthermore, this technique can be used to create hybrids between the cave and surface-dwelling populations, and thereby enable the study of the genetic basis of phenotypic adaptations to different environments.
墨西哥丽脂鲤正成为生物科学多个研究领域的模式生物。这种硬骨鱼物种近期成功的部分原因在于它拥有可杂交繁殖的洞穴种群和河流种群。这使得对这些种群在适应不同环境过程中固定下来的可遗传性状进行基因定位成为可能。虽然该物种可以在实验室中饲养和繁殖，但在白天获取胚胎以及培育品系间的杂交胚胎都具有挑战性。体外受精（IVF）已被用于多种不同的模式生物，以便在实验室中成功且重复地繁殖动物。在本方案中，我们展示了如何通过使墨西哥丽脂鲤适应不同的光照周期并结合水温变化，将繁殖周期调整到一天中的特定时间。随后，我们展示了如何识别合适的亲本鱼，从雄性和雌性亲本收集健康的配子，并使用体外受精产生有活力的后代。这使得诸如注射基因构建体或进行发育分析等相关程序能够在正常工作时间进行。此外，该技术可用于培育洞穴种群和地表种群之间的杂交种，从而能够研究表型适应不同环境的遗传基础。
