Cephalotaxus oliveri is an endemic conifer of China, which has medicinal and ornamental value. However, the limited molecular markers and genetic information are insufficient for further genetic studies of this species. In this study, we characterized and developed the EST-SSRs from transcriptome sequences for the first time. The results showed that a total of 5089 SSRs were identified from 36446 unigenes with a density of one SSR per 11.1 kb. The most common type was trinucleotide repeats, excluding mononucleotide repeats, followed by dinucleotide repeats. AAG/CTT and AT/AT exhibited the highest frequency in the trinucleotide and dinucleotide repeats, respectively. Of the identified SSRs, 671, 1125, and 1958 SSRs were located in CDS, 3′UTR, and 5′UTR, respectively. Functional annotation showed that the SSR-containing unigenes were involved in growth and development with various biological functions. Among successfully designed primer pairs, 238 primer pairs were randomly selected for amplification and validation of EST-SSR markers and 47 primer pairs were identified as polymorphic. Finally, 28 high-polymorphic primers were used for genetic analysis and revealed a moderate level of genetic diversity. Seven natural C. oliveri sampling sites were divided into two genetic groups. Furthermore, the 28 EST-SSRs had 96.43, 71.43, and 78.57% of transferability rate in Cephalotaxus fortune, Ametotaxus argotaenia, and Pseudotaxus chienii, respectively. These markers developed in this study lay the foundation for further genetic and adaptive evolution studies in C. oliveri and related species.

This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

Having a comprehensive understanding of genetic differentiation, responses to environmental change and demographic history is critical for genetic improvement and conservation efforts. Forest trees are an excellent resource for understanding population differentiation and adaptive genetic variation due to their ability to adapt to different climates and environments. Cephalotaxus oliveri is a relict conifer endemic to China. In this study, we generated transcriptome data and identified 17 728 high‐quality single‐nucleotide polymorphisms (SNPs) from 18 populations. We found significant negative correlations between expression diversity and nucleotide diversity within and among populations, suggesting that gene expression and nucleotide diversity have a reciprocal relationship when the species adapts to the environment. The analyses of population structure showed that C. oliveri displayed a striking genetic structure with four groups. BayeScEnv and RDA methods detected the signatures of local adaptation, and identified that 738 outlier SNPs were associated with precipitation, temperature and soil conditions across heterogeneous environmental conditions. Approximate Bayesian computation analyses showed that the first and second divergence occurred in the late Miocene (c. 10.075 million years ago [Ma]) and the middle Pleistocene transition (c. 0.815 Ma), respectively. Ecological niche modeling of C. oliveri revealed signs of westward expansion after the last glacial maximum, while it was predicted to experience significant range contractions in future climate change scenarios. Geographical factors and environmental factors in southern China have played a critical role in establishing the current genetic diversity and population structure of C. oliveri. This study provides an important reference for forest resource management and conservation for C. oliveri.