Amomum tsao-ko Crevost & Lemarié is an important crop that has been widely used in traditional Chinese medicine and daily diets for a long time. In this study, the genetic diversity and relationships of eight cultivated populations of A. tsao-ko grown in Southwest China were examined using sequencerelated amplified polymorphism (SRAP) and intersimple sequence repeat (ISSR) markers. The results showed that 139 (99.29%) of 140 and 185 (99.46%) of 186 bands were polymorphic by SRAP and ISSR primers amplification, respectively. The polymorphic information content of detected bands were 0.270 (SRAP) and 0.232 (ISSR), respectively. The average Nei's gene diversity (H = 0.217) and Shannon's information index (I = 0.348) at the species level generated by SRAP primer were higher than those by ISSR analysis (H = 0.158, I = 0.272). Genetic differentiation coefficients and molecular variance analysis (AMOVA) indicated that the genetic variance of A. tsao-ko mainly occurred within populations rather than among populations. The high genetic identity among populations was revealed by SRAP (0.937) and ISSR (0.963). Using UPGMA cluster analysis, principal coordinate analysis, and population structure analysis, the accessions were categorized into two major groups. Overall, results obtained here will be useful for A. tsao-ko germplasm characterization, conservation, and utilization.

Amomum tsao-ko (Zingiberaceae) is a traditional Chinese medicine and condiment, and an important economic crop in the tropical forest of southwest China. However, few simple sequence repeat (SSR) markers are available in A. tsao-ko, which is hindering genetic research in this species. The aim of this study was to develop and characterize microsatellite markers for A. tsao-ko using restriction-site-associated DNA sequencing. A total of 115,482 microsatellites were identified using MISA software, and 13,411 SSR primer pairs were designed. 100 pairs of SSR primers were selected at random and used to evaluate polymorphisms among 4 A. tsao-ko samples. Finally, 23 pairs of SSR primers with clear bands and obvious polymorphism were selected for genetic diversity analysis of 72 A. tsao-ko accessions. The number of alleles and effective number of alleles per locus ranged from 2 to 6 and from 1.315 to 3.776, respectively. The observed heterozygosity ranged from 0.208 to 0.779, and the expected heterozygosity was from 0.239 to 0.735. The average values of the polymorphic information content were 0.454. Hardy-Weinberg equilibrium (HWE) analysis showed that 10 loci significantly deviated from HWE (P < 0.05). The pairwise F ST and genetic distance values revealed low levels of genetic differentiation and high genetic similarity among six A. tsao-ko populations. These microsatellite markers developed will provide a valuable tool for further germplasm characterization, genetic diversity, and breeding studies in A. tsao-ko.

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Abstract. Amomum tsao-ko is a commercial plant that used for various purposes in medicinal and food industries. For the present investigation, 44 germplasm samples were collected from Jinping County of Yunnan Province. Clusters analysis and 2-dimensional principal component analysis (PCA) was used to represent the genetic relations among Amomum tsaoko by using simple sequence repeat (SSR) markers. Clustering analysis clearly distinguished the samples groups. Two major clusters were formed; first (Cluster I) consisted of 34 individuals, the second (Cluster II) consisted of 10 individuals, Cluster I as the main group contained multiple sub-clusters. PCA also showed 2 groups: PCA Group 1 included 29 individuals, PCA Group 2 included 12 individuals, consistent with the results of cluster analysis. The purpose of the present investigation was to provide information on genetic relationship of Amomum tsao-ko germplasm resources in main producing areas, also provide a theoretical basis for the protection and utilization of Amomum tsao-ko resources.