Liquidambar formosana (Hamamelidaceae) is a relatively fast-growing deciduous tree of high ornamental value that is indigenous to China. However, few molecular markers are available for the species or its close relatives; this has hindered genomic and genetic studies. Here, we develop a series of transferable expressed sequence tag-simple sequence repeats (EST-SSRs) for genomic analysis of L. formosana. We downloaded the sequence of the L. formosana transcriptome from the National Center of Biotechnology Information Database and identified SSR loci in the Unigene library. We found 3284 EST-SSRs by mining 34,491 assembled unigenes. We synthesized 100 random primer pairs for validation of eight L. formosana individuals; of the 100 pairs, 32 were polymorphic. We successfully transferred 12 EST-SSR markers across three related Liquidambar species; the markers exhibited excellent cross-species transferability and will facilitate genetic studies and breeding of Liquidambar. A total of 72 clones of three Liquidambar species were uniquely divided into three main clusters; principal coordinate analysis (PCoA) supported this division. Additionally, a set of 20 SSR markers that did not exhibit nonspecific amplification were used to genotype more than 53 L. formosana trees. The mean number of alleles (Na) was 5.75 and the average polymorphism information content (PIC) was 0.578, which was higher than that of the natural L. formosana population (0.390). In other words, the genetic diversity of the plus L. formosana population increased, but excellent phenotypic features were maintained. The primers will be valuable for genomic mapping, germplasm characterization, gene tagging, and further genetic studies. Analyses of genetic diversity in L. formosana will provide a basis for efficient application of genetic materials and rational management of L. formosana breeding programs.

A high-density genetic linkage map is essential for plant genetics and genomics research. However, due to the deficiency of genomic data and high-quality molecular markers, no genetic map has been published for Prince Rupprecht’s larch (Larix principis-rupprechtii Mayr), a conifer species with high ecological and commercial value in northern China. In this study, 145 F1 progeny individuals from an intraspecific cross between two elite clones of L. principis-rupprechtii and their parents were employed to construct the first genetic map in this important tree species using specific-locus amplified fragment sequencing (SLAF-seq). After preprocessing, the procedure yielded 300.20 Gb of raw data containing 1501.22 M pair-end reads. A total of 324,352 SNP markers were detected and 122,785 of them were polymorphic, with a polymorphism rate of 37.86%. Ultimately, 6099 SNPs were organized into a genetic map containing 12 linkage groups, consistent with the haploid chromosome number of larch and most other species in the Pinaceae family. The linkage map spanned 2415.58 cM and covered 99.6% of the L. principis-rupprechtii genome with an average of 0.4 cM between adjacent markers. To the best of our knowledge, this map is the first reference map for L. principis-rupprechtii, as well as the densest one obtained in larch species thus far. The genome-wide SNPs and the high-resolution genetic map will provide a foundation for future quantitative trait loci mapping, map-based cloning, marker-assisted selection, comparative genomics, and genome sequence assembly for larch trees.

Liquidambar formosana is a multipurpose tree species native to China. There has been increasing interest in L. formosana due to its leaves being rich in shikimic acid, which plays a key role in the synthesis of the antiviral drug oseltamivir phosphate. Here, shikimic acid content (SAC) and other breeding traits, including tree height (HT), diameter at breast height (DBH), height to crown base (HCB), individual tree volume (VOL), leaf color (LC) and stem straightness degree (SSD), for 387 families of 19 provenances were evaluated in a provenance–family trial of L. formosana to estimate genetic parameters and reveal geographical variation patterns and, ultimately, screen out superior provenances and families. Differences among provenances and families were significant for all tested traits, indicating a high potential for selective breeding. Broad-sense heritabilities of provenance ($h_p^2$ = 0.19–0.57) and family ($h_f^2$ = 0.16–0.31) were moderate for most traits. Moderate to strong genetic correlations were found among HT, DBH, VOL, HCB and LC (r_A = 0.339–0.982), while adverse correlations (r_A = −0.494 to −0.816) were observed between SAC and growth traits. All target traits, excluding SSD, exhibited clinal variation in response to latitudinal gradients, and a clustering heatmap divided the 19 provenances into three groups. For single-trait selection, SAC persistently had the highest genetic gains (85.14%–163.57%). A weighted index based on breeding values was used to concomitantly improve SAC, HT and DBH. At a selection rate of 25%, the genetic gains at the provenance and family levels for SAC were 36.42% and 73.52%, and those for core growth traits ranged from −2.29% to 3.49% and 4.05% to 4.47%, respectively. As far as we know, this is the first study in L. formosana to explore the inheritance of SAC and its correlations with other traditional breeding traits. The genetic parameter estimations contribute to a better understanding of the genetic basis of SAC, and the superior provenances and families obtained lay a material foundation for the development of new varieties rich in shikimic acid, thereby promoting the in-depth exploitation and utilization of germplasm resources of L. formosana.

To evaluate the efficacy of the gene-deletor system in aspen, we evaluated the system for foreign gene removal in a hybrid aspen clone, INRA 353-53 (Populus tremula × P. tremuloides). The recombinase flipping DNA (FLP) gene was under the control of the heat-inducible promoter of Gmhsp17.6-L, and the β-glucuronidase (gusA) gene which was under the control of the 35S promoter and were constructed using the gene-deletor system in the pCaLFGmFNLFG vector. Six transgenic plants and their sublines were heated at 42 °C for 8 h and gene deletion was verified by polymerase chain reaction (PCR). Three lines exhibited partial transgene deletion while the remaining three lines did not delete. Transgenic lines were evaluated by Southern-blot analyses, verifying that the six transgenic plant lines all had a single copy of transfer DNA (t-DNA). Two partial-deletion lines and two non-deletion lines were analysed for methylation and expression of promoter and recombinase. Hardly any methylation was detected in the Gmhsp17.6-L promoter or recombinase FLP gene sequences, however, the expression of the promoter and recombinase was increased significantly in the partial-deletion compared with the non-deletion line after heat-shock treatment. These results suggest that the excision efficiency had no direct relationship with methylation status of the Gmhsp17.6-L promoter and FLP recombinase, yet was affected by the expression of the Gmhsp17.6-L and FLP after heat-shock treatment.

Liquidambar formosana, endemic to China, is a multifunctional tree species valued for its wood production, urban landscaping, and medicinal applications. Here, 111 superior L. formosana accessions were genotyped using 24 novel expressed sequence tag-simple sequence repeat (EST-SSR) markers to assess genetic diversity and structure, establish DNA fingerprints, and construct a core collection. A high degree of genetic diversity was detected in the tested accessions, with mean values for the number of observed alleles (Na), polymorphism information content (PIC), and Shannon’s information index (I) recorded at 8.458, 0.579, and 1.336 per locus, respectively. Cluster analysis, principal coordinate analysis (PCoA), and population structure analysis collectively categorized these accessions into two major groups. Specifically, those from the SangZ provenance formed a distinct group, whereas accessions from other provenances exhibiting extensive gene exchange were assigned to the second group. The combined values of the probability of identity (PI) and the probability of identity among siblings (PIsibs) across 24 SSR loci were 1.475 × 10⁻¹⁹ and 2.561 × 10⁻⁸, respectively, indicating a strong ability for fingerprint identification. Unique fingerprints for the 111 accessions were established using four selected core markers. A final core collection consisting of 34 accessions was constructed using the allele maximization (M) strategy, accounting for 30.63% of the analyzed accessions. No significant differences in genetic diversity indicators, allele frequency distributions, and accession dispersion patterns were observed between the core and original collections, suggesting that the core collection could effectively represent the entire collection. This work will promote the identification, management, and conservation of L. formosana germplasm resources while providing valuable materials for the subsequent selection and breeding of this tree species.