Oligo-FISH barcode chromosome identification system provides novel insights into the natural chromosome aberrations propensity in the autotetraploid cultivated alfalfa

Abstract

Alfalfa is one of the most economically valuable forage crops in the world. However, molecular cytogenetic studies in alfalfa lag far behind other cash crops and have reached a bottleneck. Here, we developed a novel chromosome identification system by designing 21 oligo probes in specific regions of each chromosome, which can be used as a barcode to simultaneously distinguish all chromosomes in a cell. Using this system, we revealed the chromosome karyotype features and evolutionary differences among 10 cultivated alfalfa varieties. Interestingly, we also found two chromosomal variation types, i.e. aneuploidy and large chromosomal segment deletions in the seeds of three alfalfa varieties. Variation frequency analysis showed that only 7/173 seeds in those three alfalfa varieties had chromosome aberrations, which indicated that the inheritance and meiosis of alfalfa had evolved to a relatively stable state. Remarkably, 4/7 variation seeds were chromosome 2 aberrations, suggesting that chromosome 2 appears to be more susceptible to natural chromosomal aberrations than other chromosomes during inheritance. DNA sequence variation analysis showed that the difference of presence and absence variations (PAVs) among homologous copies of chromosome 2 was larger than that of the other seven chromosomes. We suggest that such large PAV divergence among homologous copies may provide the physical basis for natural chromosome 2 aberrations propensity. Our study provides a valuable and efficient tool for alfalfa’s molecular cytogenetics and sheds new insights into the propensity for natural chromosome aberrations during autopolyploid inheritance.

Acknowledgements

This work was supported equally by Tianchi Talent Project of Xinjiang (Grant number CZ001604), High-level Talents Scientific Startup Project of Shihezi University (Grant number RCZK202362), Natural Science Foundation of China (Grant number 32400496), Science and Technology Innovation Talent Project of Bingtuan (grant number 23CB008-01), and Science and Technology Research Project of Bingtuan (grant number 2023AB070).

Author contributions

Z.M. conceived the project. H.L. and Q.X. provided seeds and funding support. Q.Z. and W.W. performed the FISH experiments. Z.M. designed the oligo-FISH probes. Y.Z., Y.L., F.D., W.L., Z.Z., F.W., and H.S. participated in the data analysis. Z.M. wrote the manuscript. All authors read and approved the final manuscript.

Data availability

All data are available in the manuscript or supplementary materials.

Conflict of interest statement

The authors declare no conflicts of interest.

Supplementary Data

Supplementary data is available at Horticulture Research online.

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