稀有鮈鲫近交系微卫星多态性分析

THE APPLICATION OF MICROSATELLITE MARKERS ON GENETIC QUALITY CONTROL OF AN INBRED STRAIN OF GOBIOCYPRIS RARUS

利用17对微卫星引物对稀有鮈鲫(Gobiocypris rarus)野生群体和近交系F20和F22进行了遗传分析。结果表明在野生群体中17个微卫星位点均为多态位点,但在F20中仅有6个多态位点,F22中则仅有4个多态位点。在野生群体中共检测到64个等位基因,F20、F22分别为26、21个。近交系的平均基因纯合率均较高,其中F20为86.18%,F22达91.96%,而野生群体平均基因纯合率为46.84%。近交系平均杂合度和平均多态信息含量均较野生群体低。在近交系F20和F22中,群体间遗传相似性指数最大,其遗传距离最小,说明二者之间的亲缘关系最近。HAN系遗传多样性明显降低,已具有较高的遗传纯度。

Genetic homogeneity is the basic demand for an inbred strain of laboratory animal, which could be detected in multi-levels, including morphology, cytogenetics, biochemistry, immunogenetics, molecular genetics and so on. Biochemical markers and skin grafting were recommended methods for genetic quality control of mammalian animals in the national standard GB1492312001. However, no prescriptive methods were destined for monitoring genetic quality of aquatic laboratory animals. Gobiocypris rarus is an endemic cyprinid fish in China, distributed only in Sichuan Province. Because this species has many attractive attributes, including sensitivity to chemicals, small size, eurythermal, easily to be cared in laboratory, short life cycle （ about 4 months） and so on, it has been widely used in many. researches of fish pathology, genetics, toxicology, embryology, and physiology in China. An inbred strain of Gobiocypris rarus, named HAN strain, has been established by brother-sister mating to the 23rd generation in the laboratory since 1990. In order to investigate the genetic background of the HAN strain and to monitor its genetic quality, we examined the inbred strain on external morphometric and meristie characters, skeletal morphometries, survival of scale transplantation, electropherogram of isozymes and microsatellite polymorphism. The present study was a part of these series works on the examination of molecular genetic homogeneity： A total of 17 microsatellite markers were used to analyze on 30 individuals of F22, 20 individuals of F20 of the HAN strain in contrast with 30 individuals of wild type which were caught in Hanyuan County in 2006. All the 17 microsatellite markers exhibited to be polymorphic in wild populations, yet only six of them were polymorphic in F20 and four polymorphic in F22. Overall 64 alleles were detected in the wild population, and the number of alleles per locus ranged from 2 to 6. But in HAN, only 1 or 2 alleles could be detected in each locus, while totally 26 and 21 alleles were found in F20 and F22, respectively. The average homozygosities of these microsatellite loci were 91.96% , 86. 18% and 46.84% in F22, F20 and wild population, respectively. The average heterozygosity was 0. 5744 in wild population, significantly higher than those of F22 and F20, which showed its high levels of genetic diversity. The average polymorphism information content was 0. 5252, 0. 3838 and 0. 3837 in wild population, F20 and F22, respectively. Therefore, in contrast to wild population, the high homozygosity and low heterozygosity were found in inbred HAN strain. Among all populations, the genetic similarity index between F20 and F22 was the largest one, which showed the closest genetic distance and the nearest relationship between them. The genetic distance between wild population and F20 was larger than one between F20 and F22, and smaller than one between wild population and F22. On the whole, the present study indicated that genetic diversity of inbred strain was much lower than wild population, and high genetic purity resulted from long-time inbreeding was existed in HAN strain. Microsatellite markers were sensitive and effective methods for monitoring genetic quality of laboratory fish. The HAN strain maintained by brother-sister mating should be periodically monitored by microsatellite markers for the unpurified loci needed to be homozygosis.