Population genetics analysis of Diospyrosmun A.Chev. ex Lecomte (Ebenaceae) based on EST-SSR markers derived from a novel transcriptome.

Abstract

Diospyrosmun A.Chev. ex Lecomte (Ebenaceae), a native evergreen tree in Vietnam, has important economic and ecological values. The absence of effective and reliable molecular markers has hampered the study of D.mun's genetic diversity and population structure, even though it is an endemic and endangered species. Therefore, significant enrichment of genomic resources is urgently needed to uncover and better understand the genetic architecture of D.mun. This study aims to demonstrate an efficient and reliable tool to explore the polymorphism within D.mun germplasm. It provides a valuable platform for the breeding and conservation of this species and other endangered species worldwide. The Illumina HiSeq™ 4000 sequencing technology was applied for the transcriptomic analysis, genetic differentiation and population structure of D.mun in Vietnam. In this study, the transcriptomes of D.mun were analysed using the Illumina HiSeq^TM 4000 sequencing system and a total of 5,588,615,700 base pairs were generated. De novo assembly indicated that 91,134 unigenes were generated (average length = 645.55 bp, N50 = 957 bp, Q20 = 98.08% and Q30 = 94.51%). A total of 92,798 and 21,134 unigenes had significant similarities amongst Nr and Swiss-Prot, respectively. In the GO database, 19,929 unigenes were annotated and these genes were divided into three major categories and 50 subcategories. In the KOG analysis, 18,499 unigenes were annotated and divided into 25 gene function categories. In the KEGG analysis, 12,017 unigenes were annotated. According to the related pathways involved, they could be classified into 56 subclasses. In this study, we have identified a total of 9,391 EST-SSR markers. Ten microsatellite loci were employed to assess the genetic diversity and structure of 82 adult D.mun trees across three populations in Vietnam. The results indicated moderate levels of genetic diversity with PIC = 0.77, N_A = 3.9, N_E = 2.8, Ho = 0.56 and H_E = 0.58 and the fixation index value was recorded as positive for three populations (NS, NH and CP). Genetic differentiation among populations was low (F_ST = 0.045), suggesting limited gene flow (Nm = 5.34). This result indicates gene exchange between the populations of ancient D.mun from different geographical areas and regions. The analysis of molecular variance (AMOVA) showed that high genetic variation existed within individuals (91%) compared to amongst populations (4%). Genetic structure analysis, DAPC and the NJ tree indicated that the three populations were divided into three main clusters. With this study, we provide a molecular resoureces for the breeding and conservation of D.mun.