基于微卫星标记的鼋亲子鉴定技术

PATERNITY TEST TECHNOLOGY BASED ON MICROSATELLITE MARKER ISLET

为研究鼋(Pelochelys cantorii)保种群体的遗传信息,利用鼋转录组文库,筛选出了10对微卫星引物,以此建立了基于2组各含5个微卫星位点多重PCR体系的鼋亲子鉴定技术,并应用2组微卫星多重PCR体系,通过ABI3130遗传分析仪及PopGene3.2和CERVUS软件对56只子一代鼋进行个体基因型检测和群体遗传多样性分析,结果显示,鼋子一代群体的平均等位基因数为2.3,微卫星位点的平均多态信息含量为0.3829,平均观测杂合度和期望杂合度分别为0.6305和0.4767,处中高度水平。对子代进行亲子分析,10个微卫星位点的累积排除概率分别为73.14%(NE-1P)、90.98%(NE-2P)和98%(NE-3P),能够满足亲子鉴定的需要。且4只亲本在生殖选择中表现出差异。鼋多重PCR亲子鉴定技术的建立为群体遗传多样性分析、家系鉴定管理和选择育种提供了有效的技术手段。

The Asian giant soft-shelled turtles(Pelochelys cantorii)belong to Testudines,Trionychidae,and Pelochelys.As one of the largest aquatic turtles in China,it is also an important indicator species for the health of the river ecosystem in the Pearl River Basin and even the southern part of China.Because of the long history of P.cantorii,it is of great scientific value of environmental changes and biological evolution.P.cantorii was widely distributed in Southeast Asia of China in history.However,due to unfair trade,excessive harvesting and killing of turtles and water conservancy projects,the population of P.cantorii has been greatly reduced.Currently,P.cantorii have become an extremely endangered species.There are only 13 P.cantorii in captivity in China,which have been listed as national first-class protected animals and taken seriously by scientific researchers.With the success of artificial domestication and breeding in recent 3 years,the genetic information of the preserved population is still unknown.In this paper,we use transcriptome data design trinucleotide repeat and repeat four nucleotides microsatellite primers 30000 on,the choice of 230 pairs of primer synthesis,received 10 after two screening amplification with high efficiency and high specificity of microsatellite marker,and build 2 multiple PCR system.Genotypes and genetic diversity of 56 soft-shelled islet were detected by ABI3130 genetic analyzer and PopGene3.2 and CERVUS Software.The results showed that the average number of alleles was 2.3 and the average content of polymorphic information was 0.3829.The average observed and expected heterozygosity were 0.6305 and 0.4767,respectively.Paternity analysis of 56 progenies showed that the cumulative exclusion probabilities of 10 microsatellite loci were 73.14%(NE-1P),90.98%(NE-2P)and 98%(NE-3p),which can meet the needs of paternity testing,respectively,and the four parents showed differences in reproductive selection.The analysis of the technology can help identify the captive population of genetic information,assist researchers to family management of groups.And more importantly,in the process of F3 generations of breeding in the future,the technology can be regarded as parental choice of technical means,to get healthier,higher genetic diversity,and their offspring,for the entire group of proliferation and endangered wild work to lay a solid foundation.