Characterization of Reverse Thermo-sensitive Genic Male Sterile Lines in Cold Regions of China

Yannong S, a reverse thermo-sensitive genic male sterile （TGMS） line, is sterile at high temperature and fertile at low temperature. In the present study, the fertility of Yannong S and the japonica reverse TGMS lines derived from it was in- vestigated in Harbin, Heilongjiang from 2013 to 2015. The results showed that Yan- nong S was completely male sterile in July and August （the hottest months of the year in Harbin）, and the seed setting rate of bagged panicles was 0. We also in- ferred that the critical temperature for inducing the sterility of Yannong S was higher than 28 ℃. Most of the japonica reverse TGMS lines derived from Yannong S were completely aborted in July and August, with a seed setting rate of 0. It can be con- cluded that the reverse TGMS lines derived from Yannong S are genetically stable lines that have a long period of sterility in cold regions, so they may be the ideal genetic materials for the development and in-situ seed production of japonica hybrid rice.

Marker-Assisted Breeding of Thermo-Sensitive Genic Male Sterile Line 1892S for Disease Resistance and Submergence Tolerance

Rice line 1892S is an elite thermo-sensitive genic male sterile(TGMS)line for two-line hybrid rice production.However,1892S is susceptible to rice blast,bacterial blight and submergence.Here we reported the introduction of blast resistance(R)gene Pi9,bacterial blight R gene Xa21 and submergence tolerance gene Sub1A into 1892S genetic background through backcrossing and marker-assisted selection.The improved TGMS line 31892S and its hybrids conferred disease resistance to rice blast and bacterial blight,and showed submergence tolerance for over 14 d without significant loss of viability.The sterility-fertility conversion of 31892S was similar to that of 1892S.31892S and its derived hybrid rice had similar agronomic traits and grain quality with 1892S and the control hybrid rice,respectively.The newly developed 31892S provided an improved TGMS line for two-line hybrid rice production with disease resistance to rice blast and bacterial blight,and submergence tolerance with no yield penalty or change in grain quality.

Improving Blast Resistance of a Thermo-Sensitive Genic Male Sterile Rice Line GD-8S by Molecular Marker-Assisted Selection

The broad-spectrum blast resistance gene Pi-1, from donor line BL122, was introduced into a thermo-sensitive genic male sterile rice line GD-8S, which possessed good grain quality but high susceptibility to rice blast, by using backcross breeding and molecular marker-assisted selection. Five elite improved male sterile lines, RGD8S-1, RGD8S-2, RGD8S-3, RGD8S-4 and RGD8S-5, were selected based on the results of molecular marker analysis, spikelet sterility, recovery rate of genetic background and agronomic traits. Thirty-three representative blast isolates collected from Guangdong Province, China were used to inoculate the improved lines and the original line GD-8S artificially. The resistance frequencies of the improved lines ranged from 76.47% to 100%, much higher than that of the original line GD-8S （9.09%）. On the agronomic characters, there were no significant differences between the improved lines and GD-8S except for flag leaf length and panicle number per plant. The improved lines could be used for breeding hybrid rice with high blast resistance.

Elongation of the Uppermost Internode for Changxuan 3S,a Thermo-Sensitive Genic Male Sterile Rice Line

Changxuan 3S, a thermo-sensitive genic male sterile （TGMS） rice line with eui gene, is derived from the TGMS rice line Pei＇ai 64S by irradiation with 350 Gy of ^60Co γ-ray. To elucidate the uppermost internode elongation of the TGMS line with eui gene, Changxuan 3S and its parent Pei＇ai 64S were used to study the effects of temperature on panicle exsertion. At 24℃, the uppermost internode of Changxuan 3S elongated the fastest from the 4^th day before flowering to 0 day （flowering）, being 2.1-fold as that of Pei＇ai 64S, whereas it elongated slowly during the 12^th day to the 4^th day before flowering and the 1^st to the 3^rd day after flowering. The uppermost internode of Changxuan 3S exserted from the flag leaf sheath at 22℃, 24℃ and 26℃, and the length of elongated uppermost internode increased with the decreasing temperatures. At 28℃, though the panicles of Changxuan 3S were still enclosed in the leaf sheath, the degree of panicle enclosure was significantly lower compared with Pei＇ai 64S. Cytological studies on Changxuan 3S showed that the uppermost internode elongation was attributed to the increase of cell number and cell elongation, and the latter was more significant. Moreover, the numbers of outermost and innermost parenchyma cells and the cell length of the uppermost internode reduced with the increasing temperatures.

Differentially Expressed Genes Related to Fertility Conversion in Thermo-sensitive Genic Male Sterile(TGMS) Lines of Brassica juncea

[Objective] This study was performed to screen functional genes related to the fertility conversion of thermo-sensitive genic male sterile （TGMS） lines of Brassica juncea L. [Method] A B. juncea TGMS line K121S was selected as the experimental material. The total RNAs were isolated from fertile and sterile pollens at different development stages, including mother cell stage, tetrad stage, tricellular pollen stage and maturity stage. DDRT-PCR was carried out to identify differentially expressed genes. [Result] A total of 44 differentially expressed cDNA fragments were identified with Dot blot. And seven candidate genes related to fertility conversion of K121S were screened out by BLASTN, including callose synthase gene, aldehyde dehydrogenase gene and RNA polymerase I transcription factor RRN3 gene which were differentially expressed at the transcriptional level, H＇-ATPase gene, fructose diphosphate aldolase -class I gene, teucine-rich repeat receptor-Jike serine/threonine- protein kinase gene and alkaline/neutral invertase gene, which were differentially expressed at the post-transcriptional level. [Conclusion] The results of this study will help to explain the molecular mechanism of thermo-sensitive genic male sterility of B. juncea.

Thermo-sensitive genic male sterile (TGMS) rice line developed by population improvement

In this study population improvement or random polycross plus mass selection was adopted with using N8S, Huaizao4, Xiang2B and Zaoyoul as parents. To help comprehensively improve the important characters such as combining ability, grain quality, resistance, stigma exsertion rate, etc. The stigma exsertion rate of ZhunS, an elite TGMS line, is 78.6%, 72.4% higher than that of V20A. Hybrids produced by crossing ZhunS with R402, Minhui77 and Minhui63 increased the yield by 3.8%-5.8% and decreased grain chalkiness by 82.1%-84.4% compared with those from crossing V20A with the corresponding R lines.

Characterizations and identification of the candidate gene of rice thermo-sensitive genic male sterile gene tms5 by mapping

Previous study indicated that the thermo-sensitive genic malesterile(TGMS) gene in rice was regulated by temperature.TGMS rice plays an important role in hybrid rice production,because the application of the TGMS system in two-line breeding is laborsaving,timesaving,simple,inexpensive,efficient,and eliminating the limitations of the cytoplasmic male sterility(CMS) system.'AnnongS' is the first discovered and deeply studied TGMS rice lines in China.'AnnongS-1' and 'Y58S',two derivatives of TGMS line AnnongS,were both controlled by a single recessive gene named tms5,which was genetically mapped on chromosome 2.In this study,three populations('AnnongS-1' × 'Nanjing11','Y58S' × 'Q611',and 'Y58S' × 'Guanghui122') were developed and used for the molecular fine mapping of the tms5 gene.By analyzing recombination events in the sterile individuals using a total of 125 probes covering the tms5 region,the tms5 gene was physically mapped to a 19-kb DNA fragment between two markers 4039-1 and 4039-2,which were located on the BAC clone AP004039.After the construction of the physical map between two markers 4039-1 and 4039-2,a member(ONAC023) of the NAC(NAM-ATAF-CUC-related) gene family was identified as the candidate gene of the tms5 gene.

Molecular Markers and Candidate Genes for Thermo-Sensitive Genic Male Sterile in Rice

The discovery of thermo-sensitive genic male sterility(TGMS) has led to development of a simple and highly efficient two-line breeding system. In this study, genetic analysis was conducted using three F_2 populations derived from crosses between IR68301 S, an indica TGMS rice line, and IR14632(tropical japonica), Supanburi 91062(indica) and IR67966-188-2-2-1(tropical japonica), respectively.Approximately 1:3 ratio between sterile and normal pollen of F_2 plants from the three populations revealed that TGMS is controlled by a single recessive gene. Bulked segregant analysis using simple sequence repeat(SSR) and insertion-deletion(InDel) markers were used to identify markers linked to the tms gene. The linkage analysis based on the three populations indicated that the tms locus was located on chromosome 2 covering the same area. Using IR68301S × IR14632 F_2 population, the results showed that the tms locus was located between SSR marker RM12676 and InDel marker 2gAP0050058. The genetic distance from the tms gene to these two flanking markers were 1.10 and 0.82 cM, respectively.InDel marker 2gAP004045 located between these two markers showed complete co-segregation with the TGMS phenotype. In addition, InDel marker vf0206114052 showed 2.94 cM linked to the tms gene using F_2 populations of IR68301S × Supanburi 91062. These markers are useful tool for developing new TGMS lines by marker-assisted selection. There were ten genes located between the two flanking markers RM12676 and 2gAP0050058. Using quantitative real-time PCR for expression analysis, 7 of the 10 genes showed expression in panicles, and response to temperatures. These genes could be the candidate gene controlling TGMS in IR68301S.

Plant Temperature and Its Simulation Model of Thermo-Sensitive Genic Male Sterile Rice

Plant temperature （Tp） and its relations to the microclimate of rice colony and irrigation water were studied using a thermo-sensitive genic male sterile （TGMS） rice line, Pei＇ai 64S. Significant differences in the daily change of temperature were detected between Tp and air temperature at the height of 150 cm （TA）. From 8：00 to 20：00, Tp was lower than TA, but they were similar during 21：00 to next 7：00. The maximum Tp occurred one hour earlier than the maximum TA, though they both reached the minimum at 6：00. Tp fluctuated less than TA. At the same height, during 6：00-13：00, Tp was higher than air temperature （Ta）, and Tp reached the maximum one hour earlier than Ta. During the rest time on sunny day, Tp was close to or even a little lower than Ta. On overcast day, Tp was higher than Ta in the whole day, and both maximized at the same time. In addition, Tp was regulated by solar radiation, cloudage and wind speed in daytime, and by irrigation water at night. The present study indicated that a TA of 29.6℃ was the critical point, at which Tp was increased or decreased by irrigation water. Tp and the difference between water and air temperatures showed a conic relation. Tp fluctuation was also regulated by the absorption or reflection of solar radiation by leaves during daytime and release of heat energy during nighttime. By analysis on correlation and regression simulation, two models of Tp were established.

Mechanism of Sterility and Breeding Strategies for Photoperiod/Thermo-Sensitive Genic Male Sterile Rice

To understand the male sterility mechanism of photoperiod/thermo-sensitive genic male sterile [P（T）GMS] lines in rice, the research progress on genetics of photoperiod and/or temperature sensitive genic male sterility in rice was reviewed. A new idea was proposed to explain the sterility mechanism of P（T）GMS rice. The fertility transition from sterile to fertile is the result of cooperative regulation of major-effect sterile genes with photoperiod and/or temperature sensitive genes, but not the so-called pgms gene in P（T）GMS rice. The minor-effect genes, which exhibit accumulative effect on sterility, are the important factors for the critical temperature of sterility transition. The more minor-effect genes the sterile line holds, the lower the critical temperature of sterility transition is. The critical temperature of sterility transition will be invariable if all the minor-effect genes are homozygous. The strategies for breeding P（T）GMS rice were also proposed. The selective indices of critical photoperiod and temperature for sterility transition should be set according to varietal type and ecological region. Imposing selection pressure is a key technology for breeding P（T）GMS rice with lower critical temperature for sterility, and improving the comprehensive performance of the whole traits and combining ability is vital for breeding P（T）GMS rice lines.

Characterization and fine mapping of RTMS10, a semi-dominant reverse thermo-sensitive genic male sterile locus in rice

The discovery and application of environment-sensitive genic male sterile(EGMS) rice germplasm provide an easy method for hybrid rice breeding and have made great contributions to hybrid rice production. Typically, the photoperiod-and thermosensitive GMS(P/TGMS) lines utilized in two-line hybrid systems are male sterile under long day or/and high temperature but fertile under short day or/and low temperature conditions. However, Yannong S(Yn S), a reverse TGMS(rTGMS) line, is sterile under low temperature(<29℃) and fertile under high temperature(>29.5℃). Here, we report a genetic study on the rTGMS trait in Yn S. Interestingly, the F1 plants of the cross between Yn S and a cultivar, L422, were male sterile at 22℃ and completely fertile at 27℃. Moreover, the segregation ratio of fertile and sterile individuals in Yn S/L422 F2 populations changed from 1:3.05 to 2.95:1 when the ambient temperature increased, showing that the rTGMS trait exhibits semidominance in Yn S. We further found a locus on chromosome 10, termed RTMS10, which controls the rTGMS trait in Yn S. We then finely mapped RTMS10 to a ~68 kb interval between markers ID13116 and ID1318 by Yn S/L422 BC6 F2 populations. A near iso-genic line(NIL) NL1 from the BC6 F3 generation was developed and the pollen of NL1 became abnormal from the meiosis stage under low temperature. In summary, we identified an rTGMS locus, RTMS10, and provided co-segregated markers, which could help to accelerate molecular breeding of rTGMS lines and better understand the rTGMS trait in rice.

Study on Seed Vigor of Rice Photo-ThermoSensitive Genic Male Sterile(PTGMS) Line with Low Temperature Storage

To explore the change rule of seed vigor and seed application technology of rice PTGMS lines with low temperature storage,Zhun S,Y58S,Feng S and Guangzhan 63-4S stored in the seed bank with the temperature of 8-10 ℃ and with the relative humidity of 55% for different years were used as the materials to study the properties of seed vigor,growth and development in the field, etc. The re- sults showed that the germination potential, germination rate, germination index and vigor index of rice PTGMS lines seeds showed downward trends as the time of low temperature storage prolonged,and all of the germination rates of the seeds within five years were over 60%. The conductivity in the soaked solution of seeds was significantly increased with the prolonged low temperature storage time. Seedling rate,root length,white root number,base stem width and seedling dry weight of seeds showed downward trends as the time of low temperature storage prolonged. There was no significant difference between two integral seedling qualities of seeds within 4-6 years in low temperature storage. Compared with the germination rate of low temperature storage seeds soaking for 6,12 and 24 h,the appropriate soak time of long time low temperature storage or low vigor seeds was 12 h. A serious natural aging phenomenon occurred in low temperature storage seeds after out of the storage. The germination rate of seeds placed under natural condition for 50 d decreased obviously. The duration from seeding to heading of Zhun S,Y58S and Feng S shortened as the time of low temperature storage prolonged, while the vari- ation of panicle duration of all PTGMS lines was not obvious. In conclusion, rice PTGMS line seeds with low temperature storage in 5 years could still be used nor- mally. However.seeds after low temperature storage should be sowed as soon as possible, and the soaking time of low vigor seeds should be shortened,and the possibility that the decreased duration from seeding to heading should also be con- sidered.

Inheritance of the Male Sterility in a New Photo/Thermo-Sensitive Genie Male Sterile Line B06S of Rice

The major male sterile genes in a new photo/thermo-sensitive genie male sterile (PTGMS) line B06S of rice were analyzed by the manipulation of mixture distribution theory. The results indicated that a pair of major male sterile nuclear genes with large effects were responsible for controlling the male sterility of B06S.

Preliminary Study on Inheritance of Stigma Exertion in Wheat Thermo-photo Sensitive Genic Male Sterile Line

Stigma exertion is one of the key factors for improving the outcrossing ability of wheat thermo-photo sensitive genic male sterile（TPSGMS） line. A DH population derived from K239S/K92 S was constructed to investigate the inheritance of stigma exertion. K239 S and K92 S are TPSGMS lines with higher and lower stigma exertion rates（SER）, respectively. The SERs of parents, reciprocal crosses and the DH population were evaluated for two consecutive years. The results showed that no significant difference was observed in SER between F1 s of K239S/K92 S and K92S/K239 S,implying that stigma exertion was a trait controlled by nuclear gene（s）. In the DH population, the segregation of low and high SERs fitted to a ratio of 3 ∶1 by Chisquare test, suggesting that the stigma exertion of K239 S was controlled by one pair of recessive genes. In addition, the effects of temperature and humidity on the expression of stigma exertion were also discussed.

New Temperature Sensitive Genic Male Sterile Lines with Better Outcrossing Ability for Production of Two-Line Hybrid Rice

An investigation was carried out with three newly developed temperature sensitive genic male sterile （TGMS） lines for their floral traits, seed production potential and outcrossing ability in ten cross combinations. In the TGMS lines, fertile pollens had an average diameter of 0.89 mm while the sterile pollens was with 0.02 mm diameter.TS-29-150GY produced the biggest fertile pollens with 0.92 mm and other two lines produced relatively smaller pollens with 0.91 and 0.85 ram. Pollen fertility during the fertility reversion period was an average of 60.7%. TS-29-150GY had the maximum of 66.9% spikelet fertility whereas other two lines （TNAU18S and TNAU60S） had relatively lower spikelet fertility of 27.8% and 26.7%, respectively. Average of 17.00 g of seed yield was obtained in the TGMS lines during the fertility reversion period. TS-29- 150GY had the highest value of 21.20 g of seed yield while TNAU18S and TNAU60S produced 16.6 g and 13.2 g of seed yield, respectively. The low seed production ability of these three TGMS lines was attributed only to the environmental conditions prevailing during the period. All three TGMS lines had considerable outcrossing potential of 41.2%, 24.6% and 25.0%, respectively. The cross combinations viz. TNAU18S/IET21508 （36 g/plant）, TNAU18S/IET21044 （13 g/plant）, TNAU18S/IET21009 （26.5 g/plant）, TNAU60S/CB-09-106 （26.2 g/plant）, TNAU60S/IET21009 （14 g/plant） and TS29-150-GY/DRR 3306 （39.2 g/plant） showed perfect synchronization with acceptable hybrid seed yield, indicating suitability of TGMS system under Indian condition. Based on the outcrossing related traits viz. panicle exertion, angle of glume opening, stigma length and pollen size, TNAU18S was identified as the best, followed by TS-29-150GY.

Introgression of Gene for Non-Pollen Type Thermo-Sensitive Genic Male Sterility to Thai Rice Cultivars

For the two-line hybrid rice system, pol en sterility is regulated by recessive gene that responds to temperature. The recessive gene controlling thermo-sensitive genetic male sterility （TGMS） is expressed when the plants are grown in conditions with higher or lower critical temperatures. To transfer tgms gene（s） control ing TGMS to Thai rice cultivars by backcross breeding method, a male sterile line was used as a donor parent while Thai rice cultivars ChaiNat 1, PathumThani 1, and SuphanBuri 1 were used as recurrent parents. The BC2F2 lines were developed from backcrossing and selfing. Moreover, the simple sequence repeat （SSR） markers were developed for identifying tgms gene and the linked marker was used for assisting selection in backcrossing. The identification lines were confirmed by pol en observation. The results showed the success of introgression of the tgms gene into Thai rice cultivars. These lines will be tested for combining ability and used as female parent in hybrid rice production in Thailand.

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.

小麦光温敏核雄性不育系BS366和BS1086杂种一代结实率研究

为了解小麦光温敏核雄性不育系BS366和BS1086的育性恢复状况,评估恢复系人工定向改良对其恢复能力的效果,在不育生态区对以55份定向改良的恢复系和33份非改良恢复系(常规小麦)为父本配制的杂种一代的结实率进行了分析。结果表明,BS366和BS1086杂种一代结实率变化范围为15.42%~140.34%和20.28%~119.07%(国际法),平均结实率分别为73.02%和72.21%,二者差异不显著,说明2份母本的育性恢复能力差异不显著。BS366和BS1086杂种一代结实率主要分布于40%~110%和60%~100%间,BS1086杂种一代结实率分布更为集中。BS366和BS1086与改良父本的杂种一代平均结实率分别为84.45%和78.97%,与非改良父本的杂种一代平均结实率分别为53.97%和60.95%,改良父本的平均恢复力高于非改良父本,表明定向改良有利于提高恢复系的恢复力。BS366和BS1086与相同父本的杂种一代结实率差异在-41.66%~61.93%之间,表明相同父本对不同母本的育性恢复力存在差异。BS366和BS1086与相同改良父本的杂种一代中有61.82%的结实率差异分布于-10%~20%之间,与相同非改良父本的杂种一代中有51.52%的结实率差异在-30%~0%之间,表明同一改良父本对不同母本的育性恢复力差异小于非改良父本。BS366与14份父本、BS1086与7份父本的杂种一代结实率高于对照,其中父本14YH261、14YH551、SD036与2份母本杂种一代的结实率均高于对照,说明具有可用恢复力的父本不仅数量极少,而且对不同母本的恢复力存在较大差异。杂种一代主茎穗的平均结实率极显著大于分蘖穗。因此,建议对不同母本分别开展强恢复力的恢复系筛选,并以中选强恢复系为亲本进行定向改良,有效增加杂种一代结实率达对照水平的组合数量,为获得更多强优势杂交组合奠定基础。

香型温敏核不育系皖香22S的选育研究

皖香22S是安徽省农业科学院水稻研究所以1892S为母本、湖南湘穗种业有限责任公司引进的香型籼稻亲本常香39S为父本进行杂交选育的香型温敏核不育系,2023年8月通过安徽省技术鉴定。皖香22S株高85.0 cm,平均每穗总颖花数193.5粒,千粒重24.0 g,株型紧凑,分蘖力较强,叶片直立内卷,有香味,冷水池育性鉴定在14.5 h、23.5℃、6 d条件下,花粉不育度99.95%,套袋自交结实率0.25%。皖香22S在合肥5月20日播种,播始历期为84 d,比近似品种常香39S短5 d左右,主茎叶龄13.2叶。柱头外露率和异交结实率较高。稻瘟病、稻曲病和白叶枯病抗性较好;米质较优;所配组合杂种优势强,其中皖香22S/银月丝苗在2023年的小区优势鉴定中,较丰两优4号增产9.35%,差异达极显著水平。

Solutions to Insecurity Problems in Seed Production of Two-line Hybrid Rice

The insecure problems of seed production have seriously hampered the healthy and sustainable development of two-line hybrid rice.The safety issues on seed production of two-line hybrid rice and current research situation were pointed out in this paper.The three main reasons for unsafety in seed production of twoline hybrid rice were unsuitable site selection,high critical sterility-inducing temperature and the drift of critical temperature.In this paper,strategies and measures were put forward based on many years of practice.It could minimize the risk in seed production of two-line hybrid rice by selecting dual-purpose genic male sterile line with lower critical sterility-inducing temperature and long lower temperature resistant time.Based on the climate data and climatic demands of the ＂three safe-periods＂ in seed production,a new idea for determining appropriate bases and periods for seed production was proposed by using computer technology,which solved the aimless selection of sites and periods for the seed production of two-line hybrid rice.Besides,we established a system of single plant selection and identification method and original seed propagation with cyclic cold water,which could reduce the generation number of original seed propagation in seed production and avoid the drift of critical sterility-inducing temperature.This paper improved the seed production safety in the three aspects of seed nature,seed source and seed production site.