Transcriptome-wide genetic variations in the Leucaena genus for fingerprinting and breeding

Authors: HAN, YONG - Department Of Primary Industries; ABAIR, ALEXANDER - New Mexico State University; VAN DER ZANDEN, JULIAN - Department Of Primary Industries; Nageswara-Rao, Madhugiri - Rao; VASAN, SAIPRIYAA - Murdoch University; BHOITE, ROOPALI - Department Of Primary Industries; CASTELLO, MARIECLAIRE - Department Of Primary Industries; BAILEY, DONOVAN - New Mexico State University; REVELL, CLINTON - Department Of Primary Industries; LI, CHENGDAO - Department Of Primary Industries; REAL, DANIEL - Department Of Primary Industries

Submitted to: Agronomy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/10/2024
Publication Date: 7/12/2024
Citation: Han, Y., Abair, A., Van Der Zanden, J., Nageswara Rao, M., Vasan, S.P., Bhoite, R., Castello, M., Bailey, D., Revell, C., Li, C., Real, D. 2024. Transcriptome-wide genetic variations in the Leucaena genus for fingerprinting and breeding. Agronomy. https://doi.org/10.3390/agronomy14071519.
DOI: https://doi.org/10.3390/agronomy14071519

Interpretive Summary: The Leucaena genus consists of a variety of shrub and tree species native to the New World, with a rich history of human utilization. The species Leucaena leucocephala (Lam.) de Wit. is highly valued for its ability to fix nitrogen and is widely used for erosion control, landscaping, multipurpose farming, phytoremediation, paper production, and biofuel. However, it is considered an invasive weed in over 50 countries, which limits its use. Efforts have been made to develop non-flowering or sterile Leucaena through interspecific crosses. To identify these triploids at an early stage and avoid wasting resources on non-hybrid plants, a fast and reliable method for hybrid detection is essential. In this study, genetic variants across 19 diploid and one tetraploid species of Leucaena were identified for the first time using mRNA-based transcriptome analysis. Over 3000 Leucaena plants resulting from interspecific crosses were screened for hybridity using genetic markers from the parental lines. The developed platform can rapidly identify triploids at the seedling stage, saving time and cost for breeding programs. Furthermore, the variant database can be used for future genetic diversity studies, mapping, marker-assisted selection, and breeding in Leucaena.

Technical Abstract: Leucaena is a versatile legume shrub/tree used as tropical livestock forage and in timber industries, but it is considered a high environmental weed risk due to its prolific seed production and broad environmental adaptation. Interspecific crossings between Leucaena species have been used to create non-flowering or sterile triploids that can display reduced weediness and other desirable traits for broad use in forest and agricultural settings. However, assessing the success of the hybridisation process before evaluating the sterility of putative hybrids in the target environment is advisable. Here, RNA sequencing was used to develop breeding markers for hybrid parental identification in Leucaena. RNA-seq was carried out on all 19 diploid and one tetraploid Leucaena species, and transcriptome-wide unique genetic variants were identified relative to a L. trichandra draft genome. Over 16 million single-nucleotide polymorphisms (SNPs) and 0.8 million insertions and deletions (indels) were mapped. These sequence variations can differentiate all species of Leucaena from one another, and a core set of about 75,000 variants can be genetically mapped and transformed into genotyping arrays/chips for the conduction of population genetics, diversity assessment, and genome-wide association studies in Leucaena. For genetic fingerprinting, more than 1500 variants with even allele frequencies (0.4-0.6) among all species were filtered out for marker development and testing in planta. Notably, SNPs were preferable for future testing as they are more accurate and displayed higher transferability within the genus than indels. Hybridity testing using SNP markers was also more reliable and highly consistent with field observations. The developed markers pave the way for rapid, accurate, cost-effective diversity assessment, variety identification and breeding selection in Leucaena.