基于高通量测序的厚唇裸重唇鱼微卫星分子标记筛选

Isolation and Characterization of Microsatellite Loci for Gymnodiptychus pachycheilus by Illumina HiSeq Sequencing

本研究对厚唇裸重唇鱼(Gymnodiptychus pachycheilus)基因组进行Illumina Hi Seq高通量测序,共获得85905条序列,包含567200个微卫星位点,从中筛选出15个微卫星位点,采用雅砻江新龙种群和黄河渭河种群对其多态性进行了验证。新龙种群中,15个位点的平均等位基因数为6.9(3~13),观测杂合度(Ho)为0.7124,多态信息含量(PIC)为0.6303,有8个位点显著偏离哈迪-温伯格平衡,1对位点表现出连锁。渭河种群中,15个位点的平均等位基因数为8.4(4~16),观测杂合度(Ho)为0.7195,多态信息含量(PIC)为0.6807,有7个位点显著偏离了哈迪-温伯格平衡,4对位点表现出连锁。筛选获得的15个微卫星位点多态性较高,适合用于厚唇裸重唇鱼种群遗传学研究。

[Objectives]Microsatellite markers have been widely used in the research of conservation genetics of aquatic animals.However,there is no report on microsatellite loci of Gymnodiptychus pachycheilus.[Methods]From 2014 to 2022,a total of 2 G.pachycheilus populations(56 individuals)were collected from the main stream of the middle Yalong River and Weihe River.Illumina HiSeq sequencing was used to explore microsatellite markers from the genome of G.pachycheilus.Microsatellite loci were tested using 2 wild population samples’DNA of G.pachycheilus.Null alleles were tested using the software MICRO-CHECKER version 2.2.3,genetic diversity statistics were obtained using AUTOTET.Deviations from Hardy-Weinberg equilibrium and linkage disequilibrium at each locus were calculated using GENEPOP 4.7.5,and significance values were adjusted after applying Bonferroni correction.Polymorphism information content(PIC)value was based on PIC_Calc0.6.[Results]567200 microsatellite loci were identified from 85905 individual sequence reads produced,and 15 primer pairs were designed and testified in the Yalong River Xinlong population and the Weihe River population.In the Xinlong population,the average number of observed alleles per locus was 6.87(from 3 to 13),the average observed heterozygosities and PIC were 0.7124 and 0.6303 respectively,1 pair linkage disequilibrium was detected and eight loci showed significant deviation from Hardy-Weinberg equilibrium among the loci(Table 2).In the Weihe population,the average number of observed alleles per locus was 8.4(from 4 to 16),the average observed heterozygosities and PIC were 0.7195 and 0.6807 respectively,4 pairs linkage disequilibrium was detected and 7 loci showed significant deviation from Hardy-Weinberg equilibrium among the loci(Table 3).[Conclusion]These 15 high polymorphism microsatellite loci described for G.pachycheilus will be useful for future study on phylogeography and genetic diversity studies.