SSR molecular maker development based on three-generation full-length transcriptome sequencing of Platypharodon extremus

【Objective】The purpose of the study was to develop SSR molecular markers for Platypharodon extremus based on three-generation full-length transcriptome sequencing data, in order to accumulate and enrich the genetic information of this species,and to provide theoretical reference for guiding and optimizing artificial breeding techniques in the later stage. 【Method】Based on the three-generation full-length transcriptome sequencing results of P. extremus(PRJNA 792686), the distribution of SSRs was analyzed, and polymorphic SSR primers were selected. The genetic diversity of 37 P. extremus individuals in two watersheds was analyzed. 【Result】The results showed that the number of mononucleotide repeat SSRs was the largest in P. extremus, accounting for 37.87%. The dinucleotide-hexanucleotide repeat sequences accounted for 31.77%, 10.32%, 1.29%, 0.25% and 0.10% in order. The dinucleotide, trinucleotide, tetranucleotide and hexanucleotide repeat sequences were predominant at 5-8 repeats, accounting for 55.26%, 94.90%, 70.92% and 89.52%of the total number of their respective SSRs, respectively. After that, they gradually decreased with the increase of repeat number, and they were the lowest when the number of repeats was 21-24, accounting for 2.63%, 0.04%, 2.68% and 0 of the total number of their respective SSRs, respectively. The mononucleotide repeat sequences were predominantly repeated 9-12 times, accounting for 71.43% of their total SSRs.The pentanucleotide repeats were mainly concentrated in 5-8times, accounting for 83.33% of their total SSRs. The top 5 sequences in trinucleotide, tetranucleotide and pentanucleotide repeats were TA(17.07%),AT(16.95%), TG(15.29%), AC(12.51%)and CA(9.71%); TTA(7.53%),GAT(6.07%), TAT(5.93%), CAG(5.50%)and GAG(5.50%); TATC(4.83%), TTTA(4.32%), TTCT(3.81%), TATT(3.56%) and TGTT(3.18%), respectively. A total of 38 pairs of SSR primers were designed, and 9 pairs of SSR polymorphic primers were ultimately screened. The number of alleles(Na) in the two watersheds was 3-29, in which the expected heterozygosity(He) of P. extremus in watershed A was 0.484-0.931 and the Shannon index(H') was 0.677 to 2.793. The He of the watershed B was 0.315-0.948 and H' was 0.578-3.075. 【Conclusion】P. extremus in the two watersheds have high genetic diversities. The nine polymorphic SSR molecular markers developed can be used for breeding and germplasm protection of P. extremus.