Detection of SNPs based on DNA specific-locus amplified fragment sequencing in Chinese fir (Cunninghamia lanceolata (Lamb.) Hook)

Abstract

Compared to angiosperms, conifers represent more complex genomes with larger giga-genome size. To detect large-scale single nucleotide polymorphisms (SNPs), whole genome sequencing of a conifer population is still unaffordable. In this work, we report the use of DNA specific-locus amplified fragment sequencing (SLAF-seq) for large-scale SNP detection in Chinese fir (Cunninghamia lanceolata (Lamb.) Hook), an ecological and economic important conifer in China. SLAF libraries of 18 parent clones of a Chinese fir 2.5 generation seed orchard were sequenced and a total of 117,924 SLAFs were developed. We detected 147,376 SNPs from these SLAFs; 146,231 of them represented simple nucleotide change in A/G, C/T, A/C, A/T, C/G or G/T. The most frequent SNPs occurred in C/T (34.3%), while the majority of SNPs (68.2%) belonged to transition events (A/G and C/T). Notably, all the sequenced samples had high portion (78.2-80.9%) of common SNPs indicating that the Chinese fir genomes tended to change its nucleotides at common loci. 48,406 informative SNPs were then successfully utilized to genotype the tested samples (n = 18) followed by a phylogenetic tree to clarify their genetic relationship. Furthermore, a set of very high linkage disequilibrium (0.51-1.00) were identified from these informative SNPs. In brief, our work demonstrated that SLAF-seq is an alternative and cost-effectively high-throughput approach for large-scale SNP exploitation in Chinese fir. While the obtained SNPs offer useful marker resource for further genetic and genomic studies and will be helpful for Chinese fir breeding programs.

Chinese Fir Breeding in the High-Throughput Sequencing Era: Insights from SNPs

“…Notably, the NGS-based SNP genotyping approach, common known as genotyping by sequencing (GBS), has been readily used in the forestry tree species on population genetic studies, even for conifers that have very large (16-35 Gbp or more) and complex genomes [12][13][14][15][16][17]. For conifers, an advantage of the application of GBS is that it permits high-density SNP discovery and genotyping of a population without prior knowledge of their giga-genomes and has a power to reduce the genome complexity typically by the elimination of large set of repetitive sequences that greatly reduce the next analysis effect [12,16,18]. This feature makes it more attractive for conifer tree breeders to adopt the high-throughput sequencing (HTS) platform as a straightforward tool in their molecular breeding program.Chinese fir (Cunninghamia lanceolata) is one of the major commercial conifer species in China.…”

“…Recently, Chinese fir breeders have attempted to employ molecular marker techniques to accelerate their breeding. PCR and electrophoresis-based molecular markers including RAPD, AFLP, SRAP, ISSR, and SSR have been used for Chinese fir population genetic studies [18], while HTS technologies (e.g., NGS) open up a new perspective for genome-wide screening of single nucleotide variations (SNPs) for this species.Previously, we reported a high-throughput GBS technique, named as specific-locus amplified fragment sequencing (SLAF-seq), as being suitable for large-scale SNP exploitation in Chinese fir [18]. In this work, we extended the initial development of genome-wide SNPs in a Chinese fir advanced breeding population with 221 genotypes by using SLAF-seq, and performed a genetic analysis on this population based on the exploited SNP set.…”

“…Recently, Chinese fir breeders have attempted to employ molecular marker techniques to accelerate their breeding. PCR and electrophoresis-based molecular markers including RAPD, AFLP, SRAP, ISSR, and SSR have been used for Chinese fir population genetic studies [18], while HTS technologies (e.g., NGS) open up a new perspective for genome-wide screening of single nucleotide variations (SNPs) for this species.…”

“…Previously, we reported a high-throughput GBS technique, named as specific-locus amplified fragment sequencing (SLAF-seq), as being suitable for large-scale SNP exploitation in Chinese fir [18]. In this work, we extended the initial development of genome-wide SNPs in a Chinese fir advanced breeding population with 221 genotypes by using SLAF-seq, and performed a genetic analysis on this population based on the exploited SNP set.…”

A 3K Axiom®SNP array from a transcriptome-wide SNP resource sheds new light on the genetic diversity and structure of the iconic subtropical conifer treeAraucaria angustifolia(Bert.) Kuntze

“…[38,40–43], Cunninghamia spp. [44] and Cryptomeria japonica [45, 46]. More recently, however, with advances in technology and cost reductions, SNP arrays have become very competitive with alternative sequence based SNP genotyping methods.…”