Development of elite restoring lines by integrating blast resistance and low amylose content using MAS

Blast resistance and grain quality are major problems in hybrid rice production in China. In this study, two resistance （R） genes, Pi46 and Pita, along with the gene Wxb, which mainly affects rice endosperm amylose content （AC）, were introgressed into an elite indica restoring line, R8166, which has little blast resistance and poor grain quality through marker-assisted selection （MAS）. Eight improved lines were found to have recurrent genome recovery ratios ranging from 88.68 to 96.23%. Two improved lines, R163 and R167, were selected for subsequent studies. R167, which has the highest recovery ratio （96.23%）, showed no significant differences in multiple agronomic traits. In contrast, R163 with the lowest recovery ratio （88.68%） exhibited significant differences in heading date and yield per plant compared with the recurrent parent. At two developmental stages, R163 and R167 had greatly enhanced resistance to blast over the recurrent parent. Similar trends were also observed for agronomic traits and blast resistance in R163- and R167-derived hybrids when compared with the counterparts from R8166. In addition, R163, R167, and their derived hybrids significantly improved the grain quality traits, including amylose content （AC）, gel consistency （GC）, chalky grain rate （CGR）, and degree of endosperm chalkiness （DEC）. It confirmed the success of efficiently developing elite restoring lines using MAS in this study.

Study on the combination ability and stability of rice restore line Yihui1577

Yihui1577 was bred after over 10 generations by crossing R16 and Yunan purple rice. Many new hybrid rice varieties have been bred with Yihui1577, of which Gangyou1577. Ilyou1577, Nyou1577 and Yixiang1577 were certified by Sichuan Crop Variety Release Committee, Gangyou1577 and Yixiang1577 were certified by National Crop Variety Release Committee. The series varieties bred with Yihui1577 have been used widely with accumulated area of 1,200,000 ha. By using Yihui1577 as breeding material, an excellent restore line Yihui3003 and some other restore lines, such as Yihui1979 and Yihui315 with good quality and combination ability have been bred. Analysis of combination ability and stability of restore line Yihui1577 was made by using agricultural experimentation and statistical approach. The combination ability analysis was made by not complete dual crossing method with 4 CMS lines and 6 restorer lines. It shows that the general combination ability effect value of Yihui1577 is 0.226, while Minghui63 is -0.113. So the general combination ability of Yihui1577 is prior to that of Minghui63. Stability analysis shows that the F value of Gangyou1577 is 1.81378, while Sanyou63 is 1.40327, both doesn＇t reach significant level. So Gangyou1577 has the same stability as Sanyou63.The same conclusion is got in the stability analysis of the breeds of II you 1577, Yixiang1577, Nyou 1577 and so on.

Effect of Enhanced UV-B Radiation on Seed Setting Rate and 1 000-grain Weight of Indica Hybrid Rice Restorer Lines

[Objective] This study was to investigate the effect of enhanced UV-B radiation on seed setting rate and 1 000-grain weight of hybrid rice combinations. [Method] The seed setting rate and 1 000-grain weight of 10 new sterile indica restorer lines planted in pots under enhanced UV-B radiation and fluorescent lamps （control） were respectively measured, and the differences were compared. [Result] The enhanced UV-B radiation significantly reduced the seed setting rate of indica restorer lines, and the differences between that UV-B radiation treatment and control all reached extremely significant level. In addition, the enhanced UV-B radiation reduced the 1 000-grain weight of most indica restorer lines, and compared with that of control the difference achieved significant or very significant level. However, the effect of enhanced UV-B radiation on seed setting rate and 1 000-grain weight differed to different indica restorer lines, and the differences among restorer lines tested were significant or very significant, which indicated the possibility to screen antiUV-B radiation rice materials and combinations. Finally, the indica restorer lines 09R-14, Luhui 37 and 10R-7703 which were strongly resistant to UV-B radiation were screened out. [Conclusion] This study laid foundation for breeding hybrid rice varieties resistance to UV-B radiation.

Breeding and Application of Kehui 36, a Coldtolerant Rice Restorer Line

Kehui 36, derived from Minghui 63 / Taiyin 1（♀） // Nuo 89-1（♂）, is a new rice restorer line developed by Chongqing Normal University, which exhibits strong cold tolerance, high restoring ability, large pollen amount and other characteristics. In 2013, Kehui 36 was technically identified by Chongqing Crop Variety Appraisal Committee. Gangyou 36, a hybrid line derived from Gang 46 A / Kehui 36,was registered and released for commercial production by Chongqing Crop Variety Appraisal Committee in 2012.

Identification of a Cytoplasmic Male Sterile Line NEA in Brassica napus L.and Its Genetic Studies

Male sterile NEA plants were identified in progenies of the radiated 92P x Aggregate-silique in Brassica napus L. in 1993. Their progeny plants from test crossing and open pollination were 100% male sterile. The double-low male sterile lines JL-4 and JL-18 were bred through successive backcrossing of the double -low variety No.4 and No. 18 in Brassica napus L.to NEA. Restorer lines 6720 and 6722 with significant heterosis in F1 were developed. The results from investigating the restoring-ntaintaining relationship and inheritance of the restorer gene indicated that JL-4 and JL-18 were different from both PolCMS and Shan 2A type, and their restorer gene was controlled by a pair of dominant genie genes.

Thermal Resistance of Common Rice Maintainer and Restorer Lines to High Temperature During Flowering and Early Grain Filling Stages

Fifteen common rice maintainer lines and 26 high-yielding restorer lines were used to evaluate their thermal resistance and fertility during flowering and early grain filling stages. The rice plants were subjected to high temperature stress （39-43 ℃） for 1-15 d from main stem flowering. Based on the heat stress index, they were divided into thermal resistant lines, semi-thermal resistant lines, semi-thermal sensitive lines and thermal sensitive lines. Therefore, the maintainer lines K22B, Bobai B and V20B belonged to thermal resistant lines, whereas 11-32B, Zhongzhe B and Zhong 9B belonged to thermal sensitive lines. For rice restorer lines, Minghui 63 had the highest thermal resistance, followed by R207, P32, P929, and the lowest thermal resistant lines P62-2-2, R8006 and P51. The correlation analysis indicated that the heat stress index was significantly correlated with seed-setting rate and abortive grain rate under heat stress, but not under natural conditions. This indicated that heat stress occurred during flowering and early grain filling stages mainly decreased the seed- setting rate and significantly increased the abortive grain rate in both rice maintainer and restorer lines.

Breeding and Characterization of a New Rice Restorer Line Containing Bt Gene

Bt5198, a new rice restorer line containing Bt gene, was developed from the cross and backcross of the elite restorer line Chenghui 177 with Bt Minghui 63, a transgenic Bt restorer line. The inbred lines were evaluated using PCR amplification, test paper evaluation, insect resistance evaluation in both the laboratory and paddy fields, nursery evaluation of rice blast resistance and pedigree selection of agronomic traits. Larval mortalities on Bt5198 and Bt Minghui 63 were 100% when rice culms were inoculated with the eggs of the striped stem borer （SSB） in the laboratory. Bt5198 was highly resistant against SSB and the yellow stem borer （YSB） under field conditions. The F1 hybrids derived from Bt5198 and four cytoplasmic male sterile （CMS） lines were also highly resistant to SSB and YSB and had a significant heterosis. Two-year evaluation of rice blast resistance confirmed that the resistance levels of Bt5198 to leaf blast and neck blast were similar to those of Chenghui 177 and significantly better than those of Bt Minghui 63. Seed germination ability and pollen yield of Bt5198 were similar with Chenghui 177, suggesting that the introduction of the Bt gene into the new restorer line had no significant effects on seed vitality or the yield of seed production. To identify the presence of the Bt gene, it was effective to combine test paper examination with the evaluation of insect-resistance, both in the laboratory and under field conditions.

Genetic Analysis of Stripe Disease Resistance in Rice Restorer Line C224 Using Major Gene plus Polygene Mixed Effect Model

The inheritance of stripe disease resistance in a rice restorer line C224 was analyzed using the mixed effect model of major gene plus polygene for quantitative traits.In addition,the resistance was investigated in seven crosses of C224 with maintainer lines.The results showed that the stripe resistance of C224 was controlled by two major genes with additive-dominance-epistasis effects plus polygenes with additive-dominance effects （E-1 model）.These two genes had additive effects of-12.47% and-24.75%,respectively,showing negative dominance effects.There were significant epistasis and interaction effects between the two major genes.The heritability of the two major genes was 92.12%,while that of polygenes was 2.74%,indicating that the stripe resistance had dominant major gene effect.Of the seven crosses,five displayed high or medium resistance to the stripe disease.

Resistance Evaluation of Some Chinese Leading Rice Maintainer, Restorer lines and Their Hybrids to Magnaporthe grisea

Six isolates of Magnaporthe grisea were selected to inoculate on 10 Chinese leading maintainer lines (B-lines), 14 restorer lines (R-lines) and their F1 hybrid plants. In the tested rice materials, R-lines were proved to be more resistant to blast than B-lines. The resistance frequency of about 25% F1 hybrid plants was less than their parents. In addition, 26 isolates of M. grisea collected from different rice growing areas of China were inoculated on 13 new improved hybrid rice combinations. The resistance frequencies of 5 improved hybrids were better than those of the controls and leading varieties in rice production of China.

Indica rice restorer lines with large sink potential exhibit improved nutrient transportation to the panicle,which enhances both yield and nitrogen-use efficiency

The yield and nitrogen use efficiency(NUE)of hybrid rice combinations are closely related to restorer line.Therefore,it is essential to evaluate the agronomic characteristics of restorer lines with high yield and high NUE(HYHN).However,it is unclear which restorer lines are HYHN,and neither have the common agronomic traits of the HYHN restorer lines been identified.Aiming to address this issue,we conducted two filed experiments using three nitrogen applications,which screened five HYHN restorer lines from 15 indica restorer lines.Yield,NUE and nutrient transportation of restorer lines with different yields and NUE types were examined.Yield and total nitrogen absorption in aboveground biomass(TNA)increased,whereas NUE for grain production decreased with increasing nitrogen application levels.The HYHN restorer lines had large spikelets and high weight per panicle that were significantly positively correlated with yield and NUE.Therefore,large sink potential may be beneficial for both yield and NUE.We further studied the differences in nutrient transportation to panicles between the HYHN and low yield and low NUE(LYLN)restorer lines and found that the former had a higher nitrogen absorption level and dry matter weight ratios of panicle in maturity.Moreover,the HYHN lines also had a higher root and neck-panicle node bleeding intensity per stem after heading and more developed vascular bundles of neck-panicle nodes and leaves than the LYLN lines,which could contribute to the transportation of nutrients from root to ground and from stem and leaf to spike.Therefore,the advantages of large sink potential of the HYHN restorer lines include large nutrient accumulation in and distribution to the panicles and smooth flow of nutrients along the transportation channels.

Breeding Rice Restorer Lines with High Resistance to Bacterial Blight by Using Molecular Marker-Assisted Selection

Two bacterial blight （BB） resistance genes, Xa21 and Xa4, from IRBB24 were introduced into hybrid rice restorer line Mianhui 725, which is highly susceptible to BB, by using hybridization and molecular marker-assisted selection technology. Four homologous restorer lines were obtained through testing the R target genes with molecular markers and analyzing parental genetic background. Inoculation of the four lines and their hybrids with the specific strains of Xanthomonas oryzae pv. oryzae, P1, P6 and seven representative strains of Chinese pathotype, C Ⅰ -CⅦ, showed that all of the four lines and their hybrids were highly resistant and presented broad resistance-spectrum to BB. The hybrids of G46A / R207-2 displayed good agronomic characters and high yield potential, and R207-2 was named Shuhui 207.

Breeding of R8012,a Rice Restorer Line Resistant to Blast and Bacterial Blight Through Marker-Assisted Selection

Genetic improvement is one of the most effective strategies to prevent rice from blast and bacterial blight （BB） diseases, the two most prevalent diseases jeopardizing rice production. Rice hybrids with dural resistance to blast and BB are needed for sustainable production of food. An incomplete diallele design resulted in 25 crosses between five blast and five BB resistant germplasm accessions. Only one pair of parents, DH146 ×TM487, showed polymorphism for all the markers to identify one blast resistance gene Pi25 and three BB resistance genes, Xa21, xa13 and xa5, thus it was used in the marker-assisted selection （MAS）. F2 individuals of DH146× TM487 were genotyped using flanking markers of RM3330 and sequence tagged site （STS） marker SA7 for Pi25. The resistant F2 plants with Pi25 were used for pyramiding BB resistance genes Xa21, xa13 and xa5 identified by the markers pTA248, RM264 and RM153, respectively in subsequent generations. Finally, after selection for agronomic traits and restoration ability among 12 pyramided lines, we acquired an elite restorer line, R8012 including all four target genes （Pi25＋Xa21＋xa13＋xa5）. Hybrid Zhong 9NR8012 derived from the selected line showed stronger resistance to blast and BB, and higher grain yield than the commercial checks uniformally in experimental plots, 2007 state-wide yield trial and 2008 nation-wide yield trial. This study provides a paradigmatic example to show that MAS is a practically feasible tool in effectively pyramiding multiple resistance genes. The resultant restoring line and its hybrid would play an important role in securing rice production in China.

Analysis on the combination ability and stability of rice restore line Yihui1577

Yihui1577 was bred after over 10 generations by crossing RI6 and Yunnan purple rice. Many new hybrid rice varieties had been bred with Yihui 1577, of which Gangyou 1577, Ⅱ you1577, Nyou 1577 and Yixiang1577 were certified by Sichuan Province Crop Variety Release Committee, Gangyou1577 and Yixiang1577 were certified by National Crop Variety Release Committee. The series varieties bred with Yihui1577 had been used widely with accumulated areas of 1 200 000 hm2. By using Yihui1577 as breeding material, an excellent restore line Yihui 3003 and some other restore lines, such as Yihui 1979, Yihui 315 with good rice quality and good combination ability had been bred. Analysis of combination ability and stability about restore line Yihui 1577 was made by using agricultural experimentation and statistical approach. The combination ability analysis was made in complete dual crossing method with 4 CMS lines and 6 restorer lines. It showed that the general combination ability effect value of Yihui1577 was 0.226, while Minghui63 was -0.113. So the general combination ability ofYihui1577 was prior to that of Minghui 63. Stability analysis showed that the F value of Gangyou1577 was 1.81378, while Sanyou 63 was 1.40327, both didn＇t reach significant level. So Gangyou1577 had the same stability as Sanyou 63. The same conclusion had got in the stability analysis of the breeds of Ⅱ you 1577, Yixiang 1577, Nyou 1577 and so on.

Combining Ability of Yield Traits of Rice Restorer Lines in China and South Asia

[Objective] This study was conducted to analyze the difference in the general combining ability of yield traits of rice varieties in China and South Asia and to provide reference for the introduction and utilization of rice restorer lines from South Asia. [Method] A total of 111 hybrids were obtained by crossing 37 restorer lines in China and South Asia with Gang 46A, Lu 98A and Dexiang 074 following a NC II design. The combining ability of eight yield traits of the hybrids was analyzed in Luzhou and Deyang bases of Sichuan Province. [Result] In both bases, the eight yield traits had significant or extremely significant differences between the combina- tions, restorer lines, sterile lines or sterile line^restorer line, suggesting that there were d^erences in the genetic effects between the combinations or lines. The gen- eral combining ability （GCA） variances were all higher than special combining ability （SCA） variances in parents, which indicated that these traits were affected by gene additive effect. Restorer lines contributed more to F1 generation than male sterile liens in the eight yield traits. Nine restorer lines from India and Bangladesh were ranked in the top 15 lines with high GCA in the number of productive tillers in both bases. [Co.cl.siee] There was significant differences in the combining ability in the eight yield traits between the restorer lines from China and South Asia. The lines from India and Bangladesh were better than those from China in the GCA of the number of productive tillers, and the lines from China were better than those from India and Bangladesh in the GCA of other traits.

Breeding of Rice Restorer Line Haihui 818 of Multi-resistant Gene Pyramiding

Haihui 818 is a new restorer line by cross breeding with Hua 23 S and Huahui 8131.Good plant type,high tillering ability,fine grain quality,strong restoring ability and high yield were shown in hybrid production.After artificial inoculation and field stress and molecular market-assisted selection,the results showed that rice-blast resistance genes Pi1,BB resistance genes Xa7,and brown planthopper resistance genes Bph3,Bph14 and Bph15 were pyramided together.Among its hybrids,Bo II You 818(Bo II A/Haihui 818)was approved for commercial production by Hainan Provincial Crop Variety Approval Committee in 2015.

Breeding dominant genic male sterility restorer line of Brassica napus L.

To facilate breeding process of Brassica napus,a microspore culture and molecular marker-assisted screening combined system were proposed in this research.At early flowering stage,F1 offspring of hybridized combination HY15 A×HF06 was used as donor for microspore culture to analyze effects of colchicine concentration on embryogenic and diploid rates of microspore.Treatment with 50 mg/L colchicine resulted in embryogenic rate of3.56 embryos/bud,which was substantially higher than control(0.78 embryos/bud).A total of 1,387 embryos and 862 single plants were obtained after induction culture.Ploidy detection was performed for the regenerated plants by flow cytometry.Diploid rates of microspores treated with 50 mg/L and 70 mg/L colchicine were 17.2%and 21.0%respectively,which was significantly higher than control(10.5%).Totally 108 single plants that doubled successfully were randomly selected and screened using molecular marker BE10.Approximately 54 of108 plants generated a 305 bp amplification product,whereas the other 54 plants showed a 398 bp band,thereby satisfying 1:1 separation ratio(x0.052=0.0093).These coincided with field identification results.Findings of this study indicated that homozygous breeding material could be obtained by microspore culture in a short time,thereby remarkably accelerate breeding.

Selection of wide compatible restorer lines and their application in hybrid rice breeding

Since the middle of 1980’s, wide compatibility(WC) rice lines have been screened by ricebreeders in China and applied in hybrid ricebreeding program. Several WC lines such asPecos, T984, Lunhui 422, and 02428 withideal agronomic characters were identified. Weincorporated the WC gene into restorer linesby crossing these japonica WC lines with ob-tained indica lines. Some WC restorer lineswith indica-japonica medium type were ob-tained and their application value in intersub-specific hybrid rice breeding were evaluated. 1. Effect of crossing methods on selectionefficiencies of WC restorer lines

Inducing fertility restoration in the genic male sterile line of rice with chemical regulators

For the first time, the fertility of rice genic male sterile line was partially restored with the application of chemical regulators at Hainan Rice Breeding Nursery on Mar 1993. A single panicle of the rice plant could bear as many as 27 grains. The chemical agent that could modulate the growth of the rice plants of genic male sterile line was developed by Prof ZHOU Guangqia and his colleagues at the Biology Institute, Hunan Teachers University, China.

水稻单片段代换系对CMS-DA恢复力的鉴定和遗传分析

选用矮败型细胞质(DA-CMS)的不育系协青早A(XqA)对8个携带供体恢复基因Rf3,14个携带供体恢复基因Rf4的水稻染色体单片段代换系(Chromosome single segment substitution lines,SSSLs)和华粳籼74(HJX74,Rf3Rf3/Rf4Rf4)的恢复力进行测交鉴定,并利用恢复力最强的SSSL S18与XqA组配,采用杂交回交和分子标记辅助选择的方法,构建BC3F2群体,筛选鉴定出携带基因型Rf3Rf3/Rf4Rf4,Rf3Rf3/rf4rf4,rf3rf3/Rf4Rf4和rf3rf3/rf4rf4的BC3F2单株,观察这些植株的花粉和小穗育性,并利用213个多态性SSR标记对这些单株进行遗传背景分析,研究恢复基因Rf3和Rf4对于DA-CMS的遗传效应。结果表明:在同一细胞核背景下,22个SSSLs和HJX74对于XqA的恢复力存在着显著的不同,携带恢复基因Rf3的SSSLs恢复力均低于携带恢复基因Rf4的SSSLs,并且低于对照品种HJX74;携带Rf3的SSSL S1和SSSL S2对于XqA的花粉(小穗)育性分别为14.1%(61.7%)和15.7%(62.5%),表现出较弱的恢复力;携带Rf4的SSSL S16-SSSL S22对于XqA的花粉和小穗育性均高于87%和95%,其中SSSL S18的表现出最强的恢复力(花粉育性89.2%,小穗育性96.4%)。在构建的BC3F2群体中,携带基因型Rf3Rf3/Rf4Rf4,Rf3Rf3/rf4rf4,rf3rf3/Rf4Rf4和rf3rf3/rf4rf4单株的遗传背景片段数平均为1.0,对应于恢复基因Rf3和Rf4座位的代换片段平均长度分别为18.3 cM和12.8 cM。这4种基因型的遗传效应大小为Rf3Rf3/Rf4Rf4>rf3rf3/Rf4Rf4>Rf3Rf3/rf4rf4>rf3rf3/rf4rf4,即恢复基因Rf4的恢复力大于Rf3的恢复力,而且这两个基因表现出累积效应。

Construction of SSR Linkage Map and Analysis of QTLs for Rolled Leaf in Japonica Rice

Genetic analysis of rolled leaf is important to rice ideotype breeding. To detect loci controlling rolled leaf of japonica restorer lines, SSR marker genotypes and phenotypes of flag leaf rolling index （LRI） were investigated in Xiushui 79 （P1, a japonica rice variety）, C Bao （P2, a japonica restorer line） and 254 recombinant inbred lines derived from the cross between P1 and P2 , and in two environments. A genetic map of this cross was constructed, QTLs for LRI were detected and their interactions with environments were analyzed. Among 818 pairs of SSR primers, 90 primers showed polymorphism between P1 and P2, and 12 markers showed highly significant correlation with LRI in both environments based on single marker regression analysis. The genetic map containing 74 information loci has a total distance of 744.6 cM, with an average of 10.1 cM between two adjacent loci. Three QTLs （qRL-1, qRL-7 and qRL-8-1） were detected with two softwares： WinQTLCart 2.5 and QTLNetwork2.0. qRL-8-1 was a new locus, accounting for 15.5% and 12.8% of phenotypic variations in the two environments, respectively. The phenotypic variation explained by additive effect was 6.6%. No interaction was found between qRL-8-1 genotype and environments.