Genetic breeding and molecular identification in varieties depend on high-performance genotyping tools. The high heterozygosity of the litchi genome contributes to increased resequencing costs and elevated error rates in hybridization-based genotyping methods. In this study, a liquid chip named Litchi40K v1.0 was developed with high-depth resequencing data from 875 litchi samples, and its efficacy was validated across three different populations. In the L. chinensis var. fulvosus population, three subpopulations characterized by spatial distribution, and a total of 1110 genes were identified in the genomic regions with subpopulation differentiation. Additionally, a total of 30 significant signals associated with diverse agronomic traits were identified. The H002 haplotype of LITCHI02696, dominant in the Sub2 subgroup, significantly increased the soluble solid content in the L. chinensis var. fulvosus population. In a hybrid F1 population, a high-density genetic map was constructed and 79 dwarfing-related QTLs were identified with the liquid chip. An NAC transcription factor was identified as a candidate gene with a heterozygous frameshift variant in the male parent. To facilitate the digitization of germplasm resources, 384 SNPs were selected, and the DNA fingerprint map revealed clear genetic relationships and a total of 10 potential synonym groups or instances of bud mutations were identified in 164 main cultivated litchi varieties. This study provides cost-effective, flexible, and versatile liquid chip for genetic analysis and digitalization of germplasm resources in litchi.

Litchi chinensis var. fulvosus is an important wild litchi resource in Yunnan, China, valued for favorable agronomic traits such as early flowering, early ripening, multiple flowering cycles, and high fruit-setting ability. However, its genetic diversity and population structure remain poorly understood. In this study, 192 accessions were collected from ten counties in Yunnan Province to evaluate their geographic distribution, leaf phenotypic variation, molecular diversity, population structure, and core collection composition. Eight descriptive leaf traits, nine quantitative leaf traits, and ISSR genotyping data from seven primers were analyzed. The accessions were distributed across an altitudinal range of 169–1470 m, with clear habitat differentiation among trees of different ages. Morphological analysis revealed substantial leaf variation, with mean diversity indices of 1.19 for descriptive traits and 2.76 for quantitative traits. ISSR analysis generated 49 scorable bands, of which 34 were polymorphic, corresponding to a polymorphism rate of 68.45%. The mean Shannon–Wiener diversity index was 0.3101, indicating detectable but relatively limited molecular diversity. Integrated phenotypic and molecular analyses divided the germplasm into two subpopulations. A core collection comprising 30 accessions (about 15% of the initial population) showed the best balance between sampling efficiency and diversity retention. These results provide a practical basis for the conservation, evaluation, and efficient utilization of L. chinensis var. fulvosus genetic resources and will support breeding and genetic improvement of litchi.

The online version contains supplementary material available at 10.1186/s12870-026-08499-6.

Phyllanthus emblica L. is an important medicinal and edible fruit tree in tropical and subtropical regions, yet its population genetic structure and the ge