Microsatellite Markers and Genetic Diversity Analysis of Zacco platypus Based on Simplified Genome Sequencing
[Objective]The microsatellite molecular markers of Zacco platypus based on simplified genome sequencing(2b-RAD)were established and the genetic diversity analysis were carried out to effectively carry out the analysis of genetic diversity and molecular breeding among Z.platypus varieties,and to provide the reference for excavating and utilizing the Z.platypus germplasm resources.[Method]Genomic DNA was interrupted by the restriction endonuclease EcoRⅠ,and each sample was physically broken,and a library of 300-700 bp insert fragments was selected to obtain a large amount of genetic polymorphism tags by paired-end sequencing through the Illumina HiSeq PE150 sequencing platform.[Result]2b-RAD screening of microsatellite loci in Z.platypus obtained 41 018 SSRs with 100 bp left at each end as primers,and 1 522 SSRs with primer fragments,with a fragment primer design rate of 37.1%.The constructed libraries were of high quality,with sequencing depths meeting the typing standard at high accuracy.Among the 64 screened microsatellite loci,14 loci(21.88%)were polymorphic,and 3 loci(Zpla08,Zpla09 and Zpla10)deviated obviously from Hardy-Weinberg equilibrium due to the presence of null alleles(P<0.05),so 11 of them were selected for analyzing the genetic diversity of Z.platypus populations.Analysis of the 11 microsatellite DNA loci showed that the mean number of alleles(NA)of the seven populations of Z.platypus was 3.66,the mean Shannon's information index(I)was 0.689,the mean observed heterozygosity(HO)was 0.315,the mean expected heterozygosity(HE)was 0.354,and the mean polymorphic information content(PIC)was 0.409.[Conclusion]The development of microsatellite molecular markers for Z.platypus based on 2b-RAD can be used to assess genetic diversity,genetic structure and species identification of wild Z.platypus populations.11 of the 14 microsatellite loci in the genomic DNA of Z.platypus are moderately to highly polymorphic,and 3 loci(Zpla08,Zpla09,Zpla10)deviated from Hardy-Weinberg equilibrium and should be used with caution in population genetic studies.