Department of Mechanical Engineering, University of Minnesota, Minneapolis, MN, USA.
Center for Advanced Technologies for the Preservation of Biological Systems (ATP-Bio), University of Minnesota, Minneapolis, MN, USA.
Nat Commun. 2021 Apr 23;12(1):2412. doi: 10.1038/s41467-021-22694-z.
The development of a widely adopted cryopreservation method remains a major challenge in Drosophila research. Here we report a robust and easily implemented cryopreservation protocol of Drosophila melanogaster embryos. We present innovations for embryo permeabilization, cryoprotectant agent loading, and rewarming. We show that the protocol is broadly applicable, successfully implemented in 25 distinct strains from different sources. We demonstrate that for most strains, >50% embryos hatch and >25% of the resulting larvae develop into adults after cryopreservation. We determine that survival can be significantly improved by outcrossing to mitigate the effect of genetic background for strains with low survival after cryopreservation. We show that flies retain normal sex ratio, fertility, and original mutation after successive cryopreservation of 5 generations and 6-month storage in liquid nitrogen. Lastly, we find that non-specialists are able to use this protocol to obtain consistent results, demonstrating potential for wide adoption.
广泛采用的冷冻保存方法的开发仍然是果蝇研究中的一个主要挑战。在这里，我们报告了一种稳健且易于实施的黑腹果蝇胚胎冷冻保存方案。我们提出了胚胎通透化、冷冻保护剂加载和复温的创新方法。我们表明，该方案具有广泛的适用性，已成功应用于来自不同来源的 25 个不同品系。我们证明，对于大多数品系，超过 50%的胚胎孵化，并且在冷冻保存后，超过 25%的幼虫发育成成虫。我们确定通过杂交可以显著提高存活率，以减轻冷冻保存后存活率低的品系的遗传背景的影响。我们表明，经过 5 代连续冷冻保存和在液氮中储存 6 个月后，果蝇保持正常的性别比例、生育能力和原始突变。最后，我们发现非专业人员能够使用该方案获得一致的结果，这表明该方案具有广泛采用的潜力。
