Background: Blast disease caused by the fungal pathogen Magnaporthe oryzae is one of the big problems in rice production in China, especially for high yield hybrid varieties made from a two-line system in which thermo-sensitive genic male sterile (TGMS) lines are used. In this study, we report the introgression of a rice blast resistance gene Pi2 from VE6219 into C815S, an elite rice TGMS line, leading to the development of blast resistant TGMS lines through marker assisted selection (MAS) and phenotypic selection approaches. Results: Four new TGMS lines with blast resistance gene Pi2 were developed from C815S (an elite TGMS line susceptible to the blast, used as recurrent parent) and VE6219 (a blast resistant line harboring Pi2, used as donor parent). The pathogenicity assays inoculated with 53 blast prevalent isolates in glasshouse showed that the blast resistant frequency of the four TGMS lines was 94.3%-98.1% that is equivalent to blast resistant donor parent VE6219. The field evaluation of the new lines and hybrids made from them at a blast epidemic site also showed high resistant levels against the blast. The genetic background of the newly developed TGMS lines were examined using a whole-genome single nucleotide polymorphism (SNP) array (RICE6K) that turned out more than 83% of the genomic markers were derived from the recurrent parent. The critical temperature points of fertility-sterility alteration of the new TGMS lines were between 22°C and 23°C of daily mean temperature, which is similar to that of C815S. The complete male sterility under natural growth conditions at Wuhan last more than 80 days. Their agronomic and grain quality traits meet the requirement for two-line hybrid rice production. Conclusions: The broad-spectrum and durable rice blast resistant gene Pi2 was introgressed into the elite TGMS line C815S background. The newly developed TGMS lines can be practically used for two-line hybrid rice breeding and must play an important role in sustainable rice production in China.

Breeding two-line hybrid rice with disease resistance is an effective approach to stabilize rice yield in commercial rice production of China. We improved the blast and bacterial blight resistance of Guangzhan63-4S, an elite photoperiod- and thermo-sensitive male sterile (P/TGMS) line widely used in two-line hybrid rice, by introducing the R genes Pi2 and Xa7 conferring resistance to rice blast and bacterial blight, respectively. Through the backcrossing and gene pyramiding breeding coupled with molecular marker-assisted selection, a new P/TGMS line Hua1228S carrying Pi2, Xa7, and tms5 was developed. Based on 200,000 SNP markers by next-generation sequencing, Hua1228S covered 87.6% of the recurrent genome, as well as 4.5% of the donor genome from VE6219 and 7.9% from YR7029–39. When infected with seven tested Xanthomonas oryzae pv. oryzae strains, Hua1228S conferred high resistance (0 level) to six bacterial blight strains. Moreover, Hua1228S showed broad-spectrum resistance to rice blast isolates with a high resistance frequency of 90.91%. High levels of resistance to leaf blast and neck blast were observed under heavy disease pressure in natural field. Importantly, Hua1228S showed identical fertility-sterility alteration pattern to Guangzhan63-4S. Thus, two hybrid combinations Hua Liangyou 2821 and Hua Liangyou 284 derived from Hua1228S exhibited enhanced resistance and higher yield compared with the control variety Feng Liangyou 4. These results indicate that Hua1228S has tremendous potentiality to increase and stabilize the rice yield, through the introgression of two R genes by marker-assisted selection strategy.

Chalkiness is an important grain quality trait in rice. Chalk5, encoding a vacuolar H⁺-translocating pyrophosphatase, is a major gene affecting both the percentage of grains with chalkiness (PGWC) and chalkiness degree (DEC) in rice. Reducing its expression can decrease both PGEC and DEC. In this study, the first exon of Chalk5 was edited in the elite restorer line 9311 using the CRISPR/Cas9 system and two knockout mutants were obtained, one of which did not contain the exogenous Cas9 cassette. PGWC and DEC were both significantly reduced in both mutants, while the seed setting ratio (SSR) was also significantly decreased. Staggered sowing experiments showed that the chalkiness of the mutants was insensitive to temperature during the grain-filling stage, and the head milled rice rate (HMRR) could be improved even under high-temperature conditions. Finally, in the hybrid background, the mutants showed significantly reduced PGWC and DEC without changes in other agronomic traits. The results provide important germplasm and allele resources for breeding high-yield rice varieties with superior quality, especially for high-yield indica hybrid rice varieties with superior quality in high-temperature conditions.

Exiangsimiao (EXSM) is a high-yield and high-quality indica variety and restorer line. However, its grains are not sufficiently slender. In this study, the long-grain allele of the grain shape gene GL7/GW7 was introgressed into EXSM via marker-assisted selection combined with backcross breeding, and two improved long-grain lines (EXSM-IL1 and EXSM-IL2) were developed. Their background recovery rates reached 96.70% and 97.50%, respectively. Their grain lengths (GLs) and length-to-width ratios (LWRs) were significantly higher than EXSM. The milling quality, especially the head milled rice rate (HMRR), of both improved lines was significantly lower than EXSM. EXSM-IL1 showed no significant difference in grain yield (GY) to EXSM, although both improved lines had shorter total growth duration (TGD), longer panicle length (PL), and a lower seed setting ratio (SSR) and 1000-grain weight (KGW). Six hybrid combinations derived from the improved lines exhibited longer GL, higher LWR and narrower GW than those of the hybrid combinations derived from EXSM. The TGD and KGW of the six hybrid combinations were significantly lower compared with the hybrids from EXSM. Furthermore, the two improved lines carried 41 favorable genes associated with crucial traits. These results provide the basis for rice grain shape improvement and supply long-grain and high-quality breeding lines and restorer lines for rice production in southern China.

Achieving high yield while maintaining disease resistance is a crucial goal in rice breeding programs. In this research, two cultivated rice varieties, Jia58 and Runxiang3, were selected as parental lines. A new variety, designated as the new variety RXN2, was generated and identified through a breeding process that involved hybridization of the parental lines followed by irradiation-induced mutagenesis of the offspring. Compared with its parental lines, RXN2 shows increased plant height, higher yield, and stronger resistance to bacterial blight. Comprehensive transcriptomic and metabolic analyses indicate that pathways associated with growth, such as gibberellin and auxin signaling, are upregulated in RXN2. Meanwhile, defense-related pathways, especially those involving jasmonic acid and peroxidase metabolism, are significantly enhanced. These results provide new insights into the trade-offs between growth and defense and elucidate the genetic and metabolic underpinnings of the simultaneous improvement in grain yield and disease resistance in rice.