Objective  Populus qiongdaoensis is a plant of the Populus genus found in tropical regions of China. So far, there is few studies focus on its classification and evolution. This study aimed to understand the classification and evolution of P. qiongdaoensis in genus by third-generation full-length transcriptome sequencing and other methods.

  Method  Based on the Pacbio Sequel sequencing technology, the complete full-length transcript data of P. qiongdaoensis, P. canadensis and P. simonii under heat stress were obtained. Then, the non-synonymous substitutions (Ka), sense substitution (Ks) and Ka/Ks value of homologous genes of three Populus were calculated. We further compared the expression patterns of orthologous genes under heat stress, and constructed the phylogenetic tree of five species to analyze the genetic relationship of Populus by combining the orthologous genes of P. trichocarpa and Salix suchowensis. Finally, we cloned nuclear genes (nrDNA; UDP-SQ and POPTRDRAFT_575699) and chloroplast genes (cpDNA; atpⅠ and trnF) of P. qiongdaoensis to analyze the polymorphism of these gene sequences in the population of P. qiongdaoensis, calculated the intraspecific genetic distances of P. qiongdaoensis, and the interspecific genetic distances between P. qiongdaoensis and 19 species (five Populus groups and one out-of-class group). Based on the maximum likelihood method and the minimum evolution method, the evolutionary tree of P. qiongdaoensis and 19 tree species were constructed to analyze the genetic relationship of P. qiongdaoensis.

  Result  Third-generations of transcriptome sequencing obtained a total of 660 groups of orthologous genes from P. qiongdaoensis, P. canadensis and P. simonii. The average of Ks was 0.1505, the peak value was 0.02, and the ratio of Ka/Ks < 1 was 97.27%, showing the close relationship of the three Populus species. Analysis of orthologous gene expression patterns revealed that they had the same expression pattern under heat stress. The average genetic distance of four genes including atpⅠ, trnF, UDP-SQ and POPTRDRAFT_575699 between P. qiongdaoensis and 19 tree species was 0.011, indicating that genetic relationship between P. qiongdaoensis and poplar group was closest.

  Conclusion  The analysis of orthologous genes based on third-generation full-length transcriptome sequencing reveals that P. qiongdaoensis has a close relationship with other Populus. The results of genetic distances and constructing evolutionary trees show that P. qiongdaoensis has a close relationship with Leuce by cloning sequences of cpDNA and nrDNA genes. The polymorphism and evolutionary branch confidence of cpDNA are significantly higher than that of nrDNA, indicating that cpDNA is more capable of species discrimination than nrDNA genes in P. qiongdaoensis.