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. 2024 Oct 9;14(1):23523.
doi: 10.1038/s41598-024-75763-w.

Molecular identification and studies on genetic diversity and structure-related GC heterogeneity of Spatholobus Suberectus based on ITS2

Zi-Yi Zhao #  1 , Jia-Wen Wu #  2 , Chuan-Gui Xu  1 , You Nong  1 , Yun-Feng Huang  3 , Ke-Dao Lai  4
Affiliations

Molecular identification and studies on genetic diversity and structure-related GC heterogeneity of Spatholobus Suberectus based on ITS2

Zi-Yi Zhao et al. Sci Rep. .

Abstract

To determine the role of internal transcribed spacer 2 (ITS2) in the identification of Spatholobus suberectus and explore the genetic diversity of S. suberectus. A total of 292 ITS2s from S. suberectus and 17 other plant species were analysed. S. suberectus was clustered separately in the phylogenetic tree. The genetic distance between species was greater than that within S. suberectus. Synonymous substitution rate (Ks) analysis revealed that ITS2 diverged the most recently within S. suberectus (Ks = 0.0022). These findings suggested that ITS2 is suitable for the identification of S. suberectus. The ITS2s were divided into 8 haplotypes and 4 evolutionary branches on the basis of secondary structure, indicating that there was variation within S. suberectus. Evolutionary analysis revealed that the GC content of paired regions (pGC) was greater than that of unpaired regions (upGC), and the pGC showed a decreasing trend, whereas the upGC remained unchanged. Single-base mutation was the main cause of base pair substitution. In both the initial state and the equilibrium state, the substitution rate of GC was higher than that of AU. The increase in the GC content was partly attributed to GC-biased gene conversion (gBGC). High GC content reflected the high recombination and mutation rates of ITS2, which is the basis for species identification and genetic diversity. We characterized the sequence and structural characteristics of S. suberectus ITS2 in detail, providing a reference and basis for the identification of S. suberectus and its products, as well as the protection and utilization of wild resources.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
ITS2-based phylogenetic analysis of S. suberectus and easily confused species. The source species of different sequences are marked with different colours and shapes, and the colour of the circles at the nodes changing from green to red represent an increase in bootstrap value.
Fig. 2
Fig. 2
The intraspecific genetic distance of S. suberectus and the interspecific genetic distance between S. suberectus and other species. The red dots represent the average genetic distance.
Fig. 3
Fig. 3
Ks frequency distributions of ITS2 within S. suberectus and between S. suberectus and other species.
Fig. 4
Fig. 4
S. suberectus ITS2 haplotype network. Different haplotypes are represented by circles of different colours, and the size and number of sectors they are divided into represent the number of sequence entries that make up the haplotype. The length of the lines between haplotypes represents the number of mutation sites. Variant positions and changed bases between haplotypes are marked and connected via dashes.
Fig. 5
Fig. 5
Phylogenetic tree of S. suberectus ITS2 and consensus secondary structures of members of each clade. The colour gradient from red to green represents an increase in the degree of base conservation.
Fig. 6
Fig. 6
Comparison of the GC and equilibrium GC (GC*) contents of paired and unpaired regions of the ITS2 secondary structure. Boxplots with data points in different colours represent the GC content of paired and unpaired regions (pGC and upGC), respectively. GC* values in different regions are marked with red solid lines. The red lines for the paired and unpaired regions are marked on the right with “pGC*” and “upGC*”, respectively.
Fig. 7
Fig. 7
Base substitution rates for generating AU and GC in the initial state (I) and equilibrium state (E). Both nucleotides in the base pair were substituted to produce AU and GC. Before the substitution, they exhibited correct pairing (A) and heterozygous pairing (B), respectively. Only one nucleotide in the base pair was substituted to produce AU and GC. Before substitution, the pairs exhibited heterozygous pairing (C) and homozygous pairing (D). Different substitution processes are marked with different colours in the legend.
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