Wheat is one of the most important food crops in the world. Hybrid necrosis often occurs in the process of wheat improvement, which seriously hinders the combination of superior traits among different genotypes of wheat. In the 1960s, scientists demonstrated that the hybrid necrosis in wheat was controlled by a pair of complementary genes, Ne1 and Ne2. However, the formation mechanism of hybrid necrosis in wheat has still not been uncovered.
Recently, the research group led by Prof. LING Hongqing from the Institute of Genetics and Developmental Biology (IGDB) of the Chinese Academy of Sciences performed a series of experiments to isolate the responsible genes and illustrate the formation mechanism of wheat hybrid necrosis.
Firstly, they carried out the preliminary map of Ne1 and Ne2 by genome-wide association analysis. Then, they cloned Ne2 by map-based cloning approach using residual heterozygous lines of Ne2 locus (carrying Ne1Ne1Ne2ne2 genotype) from the recombination inbred line population derived from common wheat varieties "Zhengnong 17" (ne1ne1Ne2Ne2) and "Yangbaimai" (Ne1Ne1ne2ne2). Ne2 encodes a CC-NBS-LRR domain protein. Further, they demonstrated that Ne2 was the gene responded for wheat hybrid necrosis via knocking out of Ne2. Homozygous frameshift mutations of Ne2 generated by CRISPR/Cas9 in the genetic background of necrotic lines NIL-Ne2 and RIL-66 resulted in normal leaf growth. qPCR analysis and histological staining revealed that the immune response in the necrotic plants was continuously activated. Therefore, they concluded that up-regulated expression of Ne2 induced by Ne1 and excessive accumulation of H2O2 were closely related to the formation of hybrid necrosis in wheat.
Combined with previous reports, they speculated that Ne2 and leaf rust resistance gene LrLC10/Lr13 might be the same gene according to analysis results of genetics, collinearity of the Ne2 candidate interval and sequence alignment.
Using the developed diagnostic marker for Ne2 allele, they tested 501 common wheat materials (including 301 cultivars, 200 landraces) from different countries of the world and found that the frequency of Ne2 allele in landrace (2.0%) was much lower than that in modern cultivars (13.6%). The result showed that Ne2/LrLC10/Lr13 had been partially used during wheat genetic improvement, and its utilization was limited by the existence of Ne1 gene.
This work provides an opportunity to further investigate the molecular mechanism of hybrid necrosis, to select Lr13 by molecular marker-assisted selection, and to avoid hybrid necrosis simultaneously.
This study entitled "Ne2, a typical CC-NBS-LRR-type gene, is responsible for hybrid necrosis in wheat" was published online in New Phytologist on June 23.
This research was supported by the National Natural Science Foundation of China, the Major Basic Research Program of Shandong Natural Science Foundation, and the Research Program of Hebei Science and Technology.
Frameshift mutations of Ne2 generated by CRISPR/Cas9 in the genetic background of necrotic lines NIL-Ne2 and RIL-66 restored normal leaf growth (Image IGDB)
Contact:
Dr. LING Hongqing
Institute of Genetics and Developmental Biology, Chinese Academy of Sciences
