摘要
[Objective]The paper was to develop the microsatellite markers of Datnioides pulcher,and to screen polymorphic SSR primers.[Method]D.pulcher was performed reduced-representation genome sequencing using RAD-seq(Restriction-site associated DNA sequencing).[Result]A total of 13308806 high-quality clean reads(HQ clean reads)lengths were obtained and 26359 simple sequence repeats(SSR)loci were detected in the obtained sequence by SSR search software,which consisted of 496 repeat motifs.The most common types of repeat were A/T in single base repeat sequence,and AC/GT and A G/GT repeat units in di-base repeat sequence.AGG/CCT,AGC/CTG and AGG/CCT were the dominant types in tri-base repeat sequence.In quantity,the di-base repeat type SSR loci were the largest(19492),accounting for 73.95%of the total SSR loci.Except for mono-nucleotide type,the repetition frequency of SSR loci was mainly ranged from 4 to 9,and the number of polymorphic SSR loci was 2783.About 20066 pairs of SSR primers were successfully designed with Primer 5.0.The success rate of primer design was 76.13%.According to the repetition type and repetition number in the sequencing data,100 di-to penta-base repetition type SSRs were randomly selected and validated by PCR and polyacrylamide gel in six D.pulcher samples.A total of 73 pairs of primers were amplified by electrophoresis,and 20 of them were polymorphic.[Conclusion]The developed SSR sequences can be used for genetic analysis,genetic map construction and molecular marker assisted breeding of D.pulcher and its related species.
基金
Supported by Specialized Fund for the Basic Research Operating Expenses Program of Chinese Academy of Fisheries Sciences (2016GH07,2018GH19)
China-Asean Maritime Cooperation Fund (CAMC-2018F)
National Aquatic Germplasm Resource Sharing Platform (2019DKA30470)
