Prevalence of Ineffective Haplotypes at the Rice Blast Resistance (R) Gene Loci in Chinese Elite Hybrid Rice Varieties Revealed by Sequence-Based Molecular Diagnosis

Abstract

Multiple haplotypes at the same rice blast R-gene locus share extremely high sequence similarity, which makes the gene diagnostic method using molecular markers less effective in differentiation from one another. The composition and distribution pattern of deployed R genes/haplotypes in elite rice varieties has not been extensively analyzed. In this study, we employed PCR amplification and sequencing approach for the diagnosis of R-gene haplotypes in 54 Chinese elite rice varieties. A varied number of functional and nonfunctional haplotypes of 4 target major R-gene loci, i.e., Pi2/9, Pi5, Pik, and Pib, were deduced by referring to the reference sequences of known R genes. Functional haplotypes accounted for relatively low frequencies for the Pi2/9 (15%) and Pik (9%) loci but for relatively high frequencies for the Pi5 (50%) and Pib (54%) loci. Intriguingly, significant frequencies of 33%, 39%, 46% of non-functional haplotypes at the Pi2/9, Pik, and Pib loci, respectively, with traceable original donors were identified, suggesting that they were most likely unintentionally spread by using undesirable donors in various breeding programs. In the case of Pi5 locus, only a single haplotype, i.e., Pi5 was identified. The reactions of 54 rice varieties to the differential isolates were evaluated, which showed a good correlation to the frequency of cognate avirulence (Avr) genes or haplotypes in the differential isolates. Four R genes, i.e., Pi2, Piz-t, Pi50, and Pikm were found to contribute significantly to the resistance of the elite rice varieties. Other two genes, Pi9 and Pikh, which were not utilized in rice varieties, showed promising values in breeding durable resistance due to their high resistance frequencies to the contemporary rice blast population. The sequencebased molecular diagnosis provided a promising approach for the identification and verification of haplotypes in different R-gene loci and effective R genes valuable for breeding durable rice resistance to rice blast.

Breeding a series of new rice restorer lines with blast resistance and low‐to‐intermediate apparent amylose contents by integrating marker‐assisted and phenotypic selection

“…The gradual replacement of Wx a and Wx lv by Wx b in the female parents might be the main reason for the quality improvement of hybrid rice. In recent years, blast resistance and superior grain quality have become two of the most important goals in rice breeding in China, as the former is essential for varietal releasing in the certificating system and the latter has higher market price (Senapati et al., 2011; Xiao et al., 2020). R608, Huahui 4, R1141, 93–11 and Huazhan, as parental lines used in our breeding programme, are five elite restorer lines for hybrid rice.…”

“…From 2008 to 2021, a total of 153 hybrid varieties derived from Huazhan have been released in China (http://www.ricedata.cn/variety/). These two restorer lines were previously reported to contain multiple R genes and confer resistance to many Magnaporthe oryzae strains (Lei et al., 2013; Ma et al., 2014; Tian et al., 2016; Wu et al., 2017; Xiao et al., 2020; Yang et al., 2009). However, we found both of them showed moderately to highly susceptible reactions in some blast hotspots of Hunan province, China, in recent years, whereas Gumei 4, the donor of Pigm , displayed stable resistance to rice blast (Y.…”

Insertion of transposable elements in AVR-Pib of Magnaporthe oryzae

“…S1). The 12 novel AVR-Pib haplotypes (H01-H12) were identi ed compared to previous published alleles [3,27]. Alignment of DNA sequence assemblies of the AVR-Pib gene from 90 isolates revealed 18 mutation sites, including six mutant sites in the CDS region which weren't in the signal-peptide region (Table 2; Additional le 5 and 6: Fig.…”

“…Those results showed that the Pib gene exhibits different resistance to Chinese rice blast from different rice-growing regions. There were 42 out of 54 varieties containing Pib in Chinese elite hybrid rice varieties, two haplotypes of Pib were identi ed, and 9 different haplotypes of AVR-Pib were found among 27 M. oryzae [27]. The reactions of differential isolates on 54 rice varieties between the frequency of AVR-Pib haplotypes in the differential isolates showed a good correlation [27].…”

“…There were 42 out of 54 varieties containing Pib in Chinese elite hybrid rice varieties, two haplotypes of Pib were identi ed, and 9 different haplotypes of AVR-Pib were found among 27 M. oryzae [27]. The reactions of differential isolates on 54 rice varieties between the frequency of AVR-Pib haplotypes in the differential isolates showed a good correlation [27]. Those showed that the Pib gene widly distributed in rice varieties in China, and the adapt variation of AVR-Pib of M. oryzae are occurred.…”