Genome-wide characterization leading to simple sequence repeat (SSR) markers development in Shorea robusta

Abstract

Tropical rainforests in Southeast Asia are enriched by multifarious biota dominated by Dipterocarpaceae. In this family, Shorea robusta is an ecologically sensitive and economically important timber species whose genomic diversity and phylogeny remain understudied due to lack of datasets on genetic resources. Smattering availability of molecular markers impedes population genetic studies indicating a necessity to develop genomic databases and species-specific markers in S. robusta . Accordingly, the present study focused on fostering de novo low-depth genome sequencing, identification of reliable microsatellites markers, and their validation in various populations of S. robusta in Uttarakhand Himalayas. With 69.88 million raw reads assembled into 1,97,489 contigs (read mapped to 93.2%) and a genome size of 357.11 Mb (29 × coverage), Illumina paired-end sequencing technology arranged a library of sequence data of ~ 10 gigabases (Gb). From 57,702 microsatellite repeats, a total of 35,049 simple sequence repeat (SSR) primer pairs were developed. Afterward, among randomly selected 60 primer pairs, 50 showed successful amplification and 24 were found as polymorphic. Out of which, nine polymorphic loci were further used for genetic analysis in 16 genotypes each from three different geographical locations of Uttarakhand (India). Prominently, the average number of alleles per locus ( N a), observed heterozygosity ( H o), expected heterozygosity ( H e), and the polymorphism information content (PIC) were recorded as 2.44, 0.324, 0.277 and 0.252, respectively. The accessibility of sequence information and novel SSR markers potentially enriches the current knowledge of the genomic background for S. robusta and to be utilized in various genetic studies in species under tribe Shoreae. Supplementary Information The online version contains supplementary material available at 10.1007/s10142-023-00975-8.

Tree diversity and regeneration dynamics toward forest conservation and environmental sustainability: A case study from Nawabganj Sal Forest, Bangladesh

“…Maximum species (50%) was under 'fair' status and the same result was also found by Manna and Mishra (2017) in Lalgarh forest range of West Bengal, India. In the present study Shorea robusta showed good regeneration status which is similar to the status of Shorea robusta found in tropical moist deciduous forest in Northern India (Mishra et al, 2023) and Ranchi Sal Forest in Eastern India (Kumar & Saikia, 2020), whereas the fair regeneration status of Sal was found in Lalgarh forest range of west Bengal, India (Manna & Mishra, 2017). In the current investigation, the adult stage was absent in all species, so the condition of "no regeneration" was not observed.…”

Novel Microsatellite Loci, Cross-Species Validation of Multiplex Assays, and By-Catch Mitochondrial Genomes on Ochthebius Beetles from Supratidal Rockpools

“…NGS techniques have simplified the identification and characterization of microsatellites, establishing them as valuable markers in ecology and evolution for non-model species [64,65]. NGS techniques enabled the reliable discovery of microsatellite markers, a usual method for the description of de novo microsatellites applied in recent studies [66][67][68][69][70]. In this study, we successfully characterized new loci and mitochondrial genomes while performing optimization and cross-species validation of multiplex assays in four species of supralittoral Ochthebius.…”

An improved Shorea robusta genomic DNA extraction protocol with high PCR fidelity

“…Touchdown PCR was performed using a thermal cycler machine (Eppendorf Mastercycler Nexus). Primer SRGMS310 was used (forward sequence: 5′-ATAACCCATGTCCTGCCCAA-3′ and reverse sequence: 5′-GAATGGCCATGATTTGCCCC-3′) [ 6 ]. The amplification was performed in a 15 µl PCR reaction mixture, containing 30 ng of template DNA, 7.5 µl of 2× Taq mix (Composition: 2× Taq buffer (Cat.…”

“…Among Asian dipterocarps, predominantly in the Indian sub-continent, Shorea robusta (Vern: Sal) is an ecologically sensitive and commercially valuable woody species whose genetic diversity and phylogeny are currently uncertain due to a scarcity of data [ 6 ]. It is a prominent high-quality tree species for commercial timber production for revenue generation, as well as an outstanding supplier of construction material, tannin, gum, and oil, alongside being used as fodder and firewood [ 7 ].…”

“…Consequently, we found it necessary to develop a robust DNA extraction protocol for S. robusta that could deliver a suitable DNA concentration with high purity for routine PCR amplification. Recently, we carried out de novo low-depth genome sequencing and identification of reliable microsatellite markers in S. robusta through genomic DNA extracted with the aid of this method [ 6 ]. In the present work, we have standardized a modified high-throughput DNA extraction protocol and incorporation of an optimal concentration of a PCR additive for S. robusta that can yield relatively high-quality DNA for microsatellite markers studies.…”