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Establishment of a CRISPR/Cas9-Based Genome Editing System in Macrobrachium rosenbergii
by
Junjun Yan
1, 
Siyu Qian
1,2,
Guo Li
1,
Yujie Liu
1,
Liqian Zhou
1,3,
Tiantian Ye
1,
Cui Liu
1,
Jilun Meng
1,
Yukun Jie
1 and
Zhimin Gu
1,* 1
Xianghu Laboratory, Hangzhou 311231, China
2
College of Biological and Environmental Sciences, Zhejiang Wanli University, Ningbo 315100, China
3
School of Marine Biology and Fisheries, Hainan University, 58 People Road, Haikou 570228, China
*
Author to whom correspondence should be addressed.
Animals 2026, 16(1), 13; https://doi.org/10.3390/ani16010013 (registering DOI)
Submission received: 12 November 2025
/
Revised: 16 December 2025
/
Accepted: 18 December 2025
/
Published: 19 December 2025
(This article belongs to the Section Aquatic Animals)
Simple Summary
The giant freshwater prawn Macrobrachium rosenbergii is an important farmed species, but progress in breeding better strains has been limited because gene function is hard to study in this animal. In this work, we developed a practical method to edit genes by injecting a gene-editing tool into very early embryos. We identified how to collect fertilized eggs, the best time to inject them, and the needle design that keeps most embryos alive. Using this method, we successfully disrupted a gene needed for eye development and another that controls male traits. This system provides a foundation for understanding key genes and may eventually support the creation of improved prawn lines for aquaculture.
Abstract
The giant freshwater prawn Macrobrachium rosenbergii is a globally farmed decapod species of high economic and nutritional value, but its genetic improvement has been constrained by the lack of an efficient genome editing workflow. Here, we optimized a CRISPR/Cas9 microinjection system for targeted mutagenesis in M. rosenbergii embryos. A controlled mating-based approach enabled routine collection of one-cell embryos, and developmental staging identified 0.5–2 h post-fertilization as the optimal injection window. Optimization of the needle tip (~1 μm) minimized mechanical damage, lowering early embryo mortality to ~10%. Using this system, the eye-development gene MrPAX6 and the sex-related gene MrIAG were successfully edited. MrPAX6 editing produced a 46.9% mutation rate and an average 50% reduction in eye pigment area. MrIAG editing achieved an 84% mutation rate, while post-injection survival to hatching was approximately 35%. These results establish an efficient and reproducible CRISPR/Cas9 editing system in M. rosenbergii, providing a foundation for functional genomics and genetic improvement in this commercially important prawn species.
Keywords:
Macrobrachium rosenbergii; CRISPR/Cas9; genome editing; breeding
