In the present study, a set of ten Expressed Sequence Tag-Simple Sequence Repeats (EST-SSR) markers have been developed to contribute to the genetic studies of Pinus thunbergii. The number of alleles in the developed EST-SSR markers ranged from one to six. Observed heterozygosity was 0.000 to 0.707; the expected heterozygosity ranged from 0.000 to 0.720. Polymorphism information content (PIC) was ranged from 0.000 to 0.669. The mean of the probability of identity between two unrelated individuals (PI) and two siblings (PI-Sib) were 0.540 and 0.719, respectively. The EST-SSR markers developed in the present study, in combining with the previously developed markers, can help in understanding the genetic diversity and genetic structure of P. thunbergii populations.
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