Development of EST-SSRs based on the transcriptome of Castanopsis carlesii and cross-species transferability in other Castanopsis species.

Castanopsis carlesii (Hemsl.) Hay. is a widely distributed and dominant tree species native to subtropical China with significant ecological and economic value. Due to serious human-related disturbance, its wild resources have been increasingly reduced, and whether may result in the loss of genetic diversity. However, no population genetics studies of natural C. carlesii have been reported to date. Microsatellite markers have been a useful tool in population genetics. Therefore, we developed EST-SSR markers based on the transcriptome sequencing of C. carlesii leaves. A total of 149,380,224 clean reads were obtained, and 63,012 nonredundant unigenes with a mean length of 1,034 bp were assembled and annotated based on sequence similarity searches in the Nr, Nt, KO, SwissProt, PFAM, KOG, and GO databases. The results showed that only 5,559 (8.82%) unigenes were annotated in all seven databases, but 46,338 (73.53%) could be annotated in at least one database. A total of 31,459 potential EST-SSRs were identified in 18,690 unigenes, with an average frequency of one SSR approximately 2 kb. Among the 100 EST-SSR primer pairs designed, 49 primer pairs successfully produced the expected product by amplification, with a success rate of 49%, but only 20 primer pairs showed abundant polymorphisms. Polymorphisms were verified using 25 samples from C. carlesii in Qimen, Anhui. A total of 119 alleles were detected, with a mean number of alleles (Na) of 5.95 per locus and a mean polymorphism information content (PIC) of 0.6125. All the 20 newly developed EST-SSR markers were verified in other Castanopsis species (C. sclerophylla, C. lamontii, C. fargesii, C. eyrei and C. jucunda). Sixteen primer pairs showed successful amplification in all five Castanopsis species (80%), and the transferability ratios ranged from 90% to 100%. These developed EST-SSR markers can be applied to population genetic and germplasm evaluations of C. carlesii and related species.

Complete chloroplast genome of Castanopsis sclerophylla (Lindl.) Schott: Genome structure and comparative and phylogenetic analysis.

Castanopsis sclerophylla (Lindl.) Schott is an important species of evergreen broad-leaved tree in subtropical areas and has high ecological and economic value. However, there are few studies on its chloroplast genome. In this study, the complete chloroplast genome sequence of C. sclerophylla was determined using the Illumina HiSeq 2500 platform. The complete chloroplast genome of C. sclerophylla is 160,497 bp long, including a pair of inverted repeat (IR) regions (25,675 bp) separated by a large single-copy (LSC) region of 90,255 bp and a small single-copy (SSC) region of 18,892 bp. The overall GC content of the chloroplast genome is 36.82%. A total of 131 genes were found; of these, 111 genes are unique and annotated, including 79 protein-coding genes, 27 transfer RNA genes (tRNAs), and four ribosomal RNA genes (rRNAs). Twenty-one genes were found to be duplicated in the IR regions. Comparative analysis indicated that IR contraction might be the reason for the smaller chloroplast genome of C. sclerophylla compared to three congeneric species. Sequence analysis indicated that the LSC and SSC regions are more divergent than IR regions within Castanopsis; furthermore, greater divergence was found in noncoding regions than in coding regions. The maximum likelihood phylogenetic analysis showed that four species of the genus Castanopsis form a monophyletic clade and that C. sclerophylla is closely related to Castanopsis hainanensis with strong bootstrap values. These results not only provide a basic understanding of Castanopsis chloroplast genomes, but also illuminate Castanopsis species evolution within the Fagaceae family. Furthermore, these findings will be valuable for future studies of genetic diversity and enhance our understanding of the phylogenetic evolution of Castanopsis.

The complete chloroplast genome of Castanopsis carlesii (Hemsl.) Hay.

Castanopsis carlesii (Hemsl.) Hay. is a widely distributed and dominant tree species with significant ecological and economical values. In this study, the complete chloroplast genome sequence of C. carlesii was reported using the Illumina Hiseq 2500 platform. The complete chloroplast genome was 160,205 bp forming a typical quadripartite structure, with a pair of inverted repeated (IRs) regions of 25,670 bp, a large single copy (LSC) region of 89,849 bp, and a small single copy (SSC) region of 19,016 bp. A total of 124 functional genes were annotated, including 79 protein-coding genes, 37 tRNA genes, and eight rRNA genes. The ML phylogenetic analysis showed that the genus Castanopsis formed a clade except Castanopsis fargesii.

Moderate Genetic Diversity and Genetic Differentiation in the Relict Tree Liquidambar formosana Hance Revealed by Genic Simple Sequence Repeat Markers.

Chinese sweetgum (Liquidambar formosana) is a relatively fast-growing ecological pioneer species. It is widely used for multiple purposes. To assess the genetic diversity and genetic differentiation of the species, genic SSR markers were mined from transcriptome data for subsequent analysis of the genetic diversity and population structure of natural populations. A total of 10645 potential genic SSR loci were identified in 80482 unigenes. The average frequency was one SSR per 5.12 kb, and the dinucleotide unit was the most abundant motif. A total of 67 alleles were found, with a mean of 6.091 alleles per locus and a mean polymorphism information content of 0.390. Moreover, the species exhibited a relatively moderate level of genetic diversity (He = 0.399), with the highest was found in population XY (He = 0.469). At the regional level, the southwestern region displayed the highest genetic diversity (He = 0.435) and the largest number of private alleles (n = 5), which indicated that the Southwestern region may be the diversity hot spot of L. formosana. The AMOVA results showed that variation within populations (94.02%) was significantly higher than among populations (5.98%), which was in agreement with the coefficient of genetic differentiation (Fst = 0.076). According to the UPGMA analysis and principal coordinate analysis and confirmed by the assignment test, 25 populations could be divided into three groups, and there were different degrees of introgression among populations. No correlation was found between genetic distance and geographic distance (P > 0.05). These results provided further evidence that geographic isolation was not the primary factor leading to the moderate genetic differentiation of L. formosana. As most of the genetic diversity of L. formosana exists among individuals within a population, individual plant selection would be an effective way to use natural variation in genetic improvement programs. This would be helpful to not only protect the genetic resources but also attain effective management and exploit genetic resources.