Rapid cloning of genes in hexaploid wheat using cultivar-specific long-range chromosome assembly

Nat Biotechnol. 2017 Aug;35(8):793-796. doi: 10.1038/nbt.3877. Epub 2017 May 15.


Cereal crops such as wheat and maize have large repeat-rich genomes that make cloning of individual genes challenging. Moreover, gene order and gene sequences often differ substantially between cultivars of the same crop species. A major bottleneck for gene cloning in cereals is the generation of high-quality sequence information from a cultivar of interest. In order to accelerate gene cloning from any cropping line, we report 'targeted chromosome-based cloning via long-range assembly' (TACCA). TACCA combines lossless genome-complexity reduction via chromosome flow sorting with Chicago long-range linkage to assemble complex genomes. We applied TACCA to produce a high-quality (N50 of 9.76 Mb) de novo chromosome assembly of the wheat line CH Campala Lr22a in only 4 months. Using this assembly we cloned the broad-spectrum Lr22a leaf-rust resistance gene, using molecular marker information and ethyl methanesulfonate (EMS) mutants, and found that Lr22a encodes an intracellular immune receptor homologous to the Arabidopsis thaliana RPM1 protein.

MeSH terms

  • Biotechnology
  • Cloning, Molecular / methods*
  • Edible Grain / genetics*
  • Genes, Plant / genetics*
  • Genetic Linkage
  • Triticum / genetics*