Abstract
The Western clawed frog, Xenopus tropicalis, is a highly promising model amphibian, especially in developmental and physiological research, and as a tool for understanding disease. It was originally found in the West African rainforest belt, and was introduced to the research community in the 1990s. The major strains thus far known include the Nigerian and Ivory Coast strains. However, due to its short history as an experimental animal, the genetic relationship among the various strains has not yet been clarified, and establishment of inbred strains has not yet been achieved. Since 2003 the Institute for Amphibian Biology (IAB), Hiroshima University has maintained stocks of multiple X. tropicalis strains and conducted consecutive breeding as part of the National BioResource Project. In the present study we investigated the inbreeding ratio and genetic relationship of four inbred strains at IAB, as well as stocks from other institutions, using highly polymorphic microsatellite markers and mitochondrial haplotypes. Our results show successive reduction of heterozygosity in the genome of the IAB inbred strains. The Ivory Coast strains clearly differed from the Nigerian strains genetically, and three subgroups were identified within both the Nigerian and Ivory Coast strains. It is noteworthy that the Ivory Coast strains have an evolutionary divergent genetic background. Our results serve as a guide for the most effective use of X. tropicalis strains, and the long-term maintenance of multiple strains will contribute to further research efforts.
Highlights
Amphibian species have featured prominently as experimental animals due to several key characteristics: 1) Their skeleton and internal organs are similar to those of mammals; 2) Their ovulation is controllable, resulting in many large eggs and embryos that develop robustly; 3) They display dynamic metamorphosis from tadpoles to adult frogs and 4) They have a high regenerative capacity
Inbred strains are essential for high efficiency of advanced reverse genetic tools such as ZFN, TALEN, and CRISPR/Cas9, which have appeared in recent years [6,7,8,9,10]
The SSLP loci were found to be scattered on numerous, fragmented scaffolds in assembly 4.1, our results of ePCR showed that all loci, except for 049H07, 115G04, and 045E02, were localized on the same number of scaffolds as in assembly 7.1. 049H07 and 045E02 were not localized on any scaffold, and 045E02 of LG 3 was localized on scaffold 7
Summary
Amphibian species have featured prominently as experimental animals due to several key characteristics: 1) Their skeleton and internal organs are similar to those of mammals; 2) Their ovulation is controllable, resulting in many large eggs and embryos that develop robustly; 3) They display dynamic metamorphosis from tadpoles to adult frogs and 4) They have a high regenerative capacity. Recent determination of the full nucleotide sequence of X. tropicalis offers a unique opportunity to combine forward and reverse genetics [1,4]. These advantages are accelerating the popularity of X. tropicalis. Inbred strains are important in reducing inter-individual variation and the number of animals used in investigations, as well as in facilitating reproducibility and uniformity of experimental results, e.g. for accurate genome sequencing, and for research on immune systems and functions [5]. Inbred strains are essential for high efficiency of advanced reverse genetic tools such as ZFN, TALEN, and CRISPR/Cas, which have appeared in recent years [6,7,8,9,10]. Multiple strains with different genetic backgrounds are essential for genetic screens in forward genetics
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