Genetic Quality Control Of An Inbred Strain Of Gobiocypris Rarus

Gobiocypris rarus is an endemic cyprinid fish in China, distributed only in paddy fields, ditches or gullies in some counties of Sichuan Province, such as Hanyuan county, Pengzhou county and so on. It belongs to Cypriniformes, Cyprinidae. This species has many attactive attributes, which make it possible to become a potential candidate of experimental model fish in future in China, or even all over the world. In order to use G. rarus as an laboratory animal, an inbred strain has been established by brother-sister mating to the 20th generation in the laboratory since 1990. According to the definition of inbred strain, it is time to examine genetic quality for the inbred strain. In the present study, genetic quality of G. rarus was studied by means of the methods, which are usually used in genetic monitoring of mammal experimental animal mouse, from the following five levels:1. Morphological variability between non-inbred populations (3 wild populations and 1 F1) and inbred stocks (F20 and F22) of G. rarus was studied by multivariate analysis on morphometric and meristic characters determined by traditional method and truss network, including stepwise discriminant analysis, principal component analysis and cluster analysis included. The results showed tiny intraspecies variations on meristic characters were found, but they were not effective for population distinctions. On the contrary, analysis on morphometric characters revealed abundant variations among stocks, especially between wild populations and inbred stocks . The average discriminant accuracy for all populations was over 80%, whereas the discriminant accuracy for inbred strains was much higher. In the cluster analysis, fully and excellent classification could be obtained between the two inbred stocks F10, F20 and the other populations. In the principal component analysis and stepwise discriminant analysis, the characters related to body depth of fish played important role in the discrimination of populations. It was argued that morphological differences among stocks were resulted from both genetic differences and environmental factors. Multivariate analysis on morphometric characters could be used to distinguish inbred strains of G. rarus.2. Based on the observation of head skeleton of wild population and inbred stock F20 of G. rarus, there was obvious shape difference on operculum between different populations. In the discriminant analysis, the discrimintant accuracy of wild population and F20 was 100% and 93.8%, respectively. All the results indicated that it is feasible to differentiate populations of G. rarus by means of performing discriminate analysis on fish operculum.3. Genetic purity of inbred stock of G. rarus was evaluted by scale transplantation. The results showed that the survival rate of transplanted scales in inbred stock was lower than that of in scale autotransplantation of either inbred stock or wild population, but it was much higher than that of allotransplanted scales in wild population, indicating that genetic uniformity existed among inbred stock. So it is feasible to perform genetic quality control by means of scale transplantation.4. By the polyacrylamide gel electrophoresis, six isozymes and one muscle protein from four different tissues of G. rarus were studied, and also were analyzed in both wild population and inbred strain F22. The results showed that all isozymes presented tissue specificity. Liver can be used as the suitable material to do ADH isozyme analysis among populations, and muscle used for the others. Six isozymes presumably were encoded by 13 gene loci, 11 of which were found monomorphic in wild population. And only 2 loci with two alleles are polymorphic, being EST-2, EST-3. Mean proportion of polymorphic loci was 15.56%. However, no polymorphic loci was found in F22. In addition, no difference between individuals was found in inbred stock with regard to three belts of muscle protein, whereas belt I and II presented polymorphic in wild population. That is to say, after successive mating cross of brother and sisiter, inbred stock of G. rarus has reached a high degree of genetic purity.5. The polymorphism of 17 microsatellites in wild population and inbred strain F20 and F22 of G. rarus was analyzed. All primers showed polymorphic in wild populations, yet only six of these primers were polymorphic in F20, and four pairs of primers showed polymorphic in F22. There were totally 64 alleles of these microsatellite loci in wild population, while 26 and 21 alleles were found in F20 and F22, repectively. The mean rates of homozygote in inbred stocks was 91.96%,86.18% and 46.84% in inbred stock F22, F20 and wild population, respectively. In contrast to wild population, high homozygosity and low heterzygosity were found in inbred stocks. Among all populations, the genetic distance was the closest and genetic similarity indices was the biggest between inbred stocks F20 and F22, showing they the nearest in relationship. So it was argued that genetic diversity of inbred stock was much lower than wild populations, and high degree of inbreeding existed in inbred stocks.