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.

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.

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.

Male Sterile Lines of Zinnia elegans and Their Cytological Observations

In order to find out a new pathway for utilizing heterosis of Zinnia elegans and accelerate breeding process, the mechanism of anther development of a male sterile line was explored. Backcross, sibmating, selling of fertile plants and testcross with inbred lines were analyzed and identified in the field, and cytology was observed. Recessive nucleus male sterile line AH209AB capable of being a maintainer was obtained by successive backcrosses with male sterile plants and fertile F1 plants as male parents. Cytological and anatomical studies indicated that： （1） The wall of normal anther was constituted of four layers of cells such as epidermis, powder chamber wall, middle level and tapetum cells. The process in meiosis of pollen mother cell in Zinnia elegans was normal and cytoplasm divided simultanously. Mature pollen grain was tricellular type. （2） The petal of male sterile plant degraded as a thread-like structure, the stamens were villiform in appearance and no pollens were formed. The result showed that the anther of male sterile plant no longer proceed to differentiate spore mother cell and the pollen sac after the formation of the tissue of sporogenous cells, there was no evident boundary between tapetum cell, middle lamella and inner wall of PMC, tapetal cells did not develop from the very beginning. So the abortion type was completely structural male sterility. The male sterile line belongs to non-sporange male sterile type and is of great use in F1 seeds production.

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.

Development and Application of SCAR Markers for Discriminating Cytoplasmic Male Sterile Lines from Their Cognate Maintainer Lines in Indica Rice

The DNAfragments about 1 600 bp were amplified using random amplified polymorphism DNA （RAPD） primer OPAl2 with the templates of mitochondrial DNA of Zhenshan 97A and Zhenshan 97B, and were sequenced. The nucleotide sequences and lengths of the fragments from Zhenshan 97A and Zhenshan 97B showed no difference. The precise length of the fragment was 1 588 bp. Sequence characterized amplification region （SCAR） primers were then developed to discriminate the cytoplasmic male sterile （CMS） lines and their maintainer lines. A specific 1 588 bp fragment could be amplified with SCAR primers, CHI19F2/CHI19R2 and CHI20F3/CHI23R3, in the mitochondrial DNA of Zhenshan 97A, but not Zhenshan 97B. Furthermore, the specific fragment could be also amplified from the total DNA from green leaf tissues of Zhenshan 97A with SCAR primers, but not Zhenshan 97B. With the corresponding primers, the specific fragment could also be amplified from the total DNA of green leaves of other two CMS lines with wild abortive type cytoplasm （CMS-WA）, namely Zhenpin A and Tianfeng A, but not in their maintainer lines. Moreover, using total DNA as template, each of the four pairs of SCAR primers could also be used to amplify the 1 588 bp fragment in CMS-ID （Indonesia paddy type） line 11-32A but not in 11-32B, and the specific fragment was amplified from the DNA of both F1 and F2 seedlings of Shanyou 63. The results of detecting the genetic purity of a man-made mixture of the seeds of Zhenshan 97A using CHI20F3/CHI23R3 were completely consistent with the phenotypes. Taken together, these results indicated that the specific 1 588 bp-fragment amplified by CHI20F3/CHI23R3 was the unique amplification products of CMS mitochondrial DNA, and could be used to distinguish CMS-WA and CMS-ID lines from their corresponding maintainer lines at the seedling stage.

Molecular identification of the cytoplasmic male sterile source from Dongxiang wild rice(Oryza rufipogon Griff.)

The use of cytoplasm male sterility （CMS） is crucial for three-line hybrid seed production. Two types of CMS have been discovered from Dongxiang wild rice, namely the wild abortion type CMS （CMS-WA） and the Dongxiang wild type CMS （CMS-DW）. In this study, we show the molecular identification of the two types of CMS in Dongxiang wild rice. WA352, which conferred CMS-WA, was not detected in Dongxiang wild iice, implying Dongxiang wild rice does not carry the CMS- WA source. Further analysis of WA352 in DYIA, a CMS-DW line, by PCR amplification and sequencing, revealed two insertion-deletion polymorphisms occurred in CMS-DW compared to WA352 of CMS-WA. It was reported that WA352 was comprised of an unknown origin sequence and partial sequences of three open reading frames （ORFs）, orf284, orf224 and orf288. The 42-bp insertion was located between the two segments of orf224 and orf288, which created a new chimeric ORF, orf216. This new ORF was also detected in CMS-HL. Based on the 9-bp deletion in orf284, a specific mitochon- drial marker of DW-M1 was developed, which could be used to specifically distinguish the DW-type source. Moreover, semi-quantitative RT-PCR analysis preliminarily suggested that both orf216 and orf284 could be considered as candidates for CMS-DW. These findings present a preliminary understanding of CMS-DW at the molecular level.

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.

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.

Comparisons on Genetic Diversity among the Isonuclear-Alloplasmic Male Sterile Lines and Their Maintainer Lines in Rice

Four sets of rice isonuclear-alloplasmic lines including 16 male sterile lines and their maintainer lines were analyzed by using 91 pairs of SSR primers to study the genetic diversity of nuclear genome and their relative relationships. A total of 169 alleles were detected in the 16 lines, with a frequency of polymorphic loci of 53.85% and an average number of alleles per locus of 1.8, and the average gene diversity was 0.228. Four sets of the isonuclear-alloplasmic male sterile lines shared 146 identical alleles, corresponding to 86.39% of the total alleles; meanwhile, there are 23 different alleles among the tested materials, being 13.61% of the total alleles. On average, 78.70% identical alleles and 21.30% different alleles of the total alleles were detected between the isonuclear-alloplasmic male sterile lines and their maintainer lines. There were 53.85% identical alleles and 46.15% different alleles of the total alleles among the homozygous allonucleus male sterile lines. The fingerprints were established for some male sterile lines and maintainer lines. All the materials tested were divided into three groups at the 0.2 genetic distance based on the cluster analysis. Eight lines of Huanong A and Huayu A （including Huanong B and Huayu B） were in the first group, four lines of Kezhen A （including Kezhen B） in the second group, and four lines of Zhenshan 97A （including Zhenshan 97B） in the third group. For the isonuclear-alloplasmic male sterile lines, the similarity coefficient between Y （Yegong） type and WA （wild abortive） type or between CW （Raoping wild rice） and WA type reached 87-98%.

Plant Temperature for Sterile Alteration of a Temperature-Sensitive Genic Male Sterile Rice,Peiai64S

The forecast of sterile alteration for the temperature-sensitive genic male sterile （TGMS） line in two-line hybrid rice seed production was traditionally based on screen temperature determined by weather station. The article put forward a new approach based on plant temperature, which was more exact and direct than the traditional method. The result of the simulation of the self-seeded setting rate of a widely used TGMS line, Peiai64S, by several temperature parameters and durations, showed that the fertility was directly affected by the plant temperature at a height of 20 cm or the air temperature around it in three days duration. Using the stem temperature of three days at a height of 20 cm as the simulation parameter, the fertility of Peiai64S had the maximum, minimum and optimum temperatures as 22.8, 21.7 and 22.5℃, respectively, whereas 23.2, 21.5 and 21.8℃ when using the air temperature of three days around the height of 20 cm as the parameter. Such temperature indices can be used to conclude the sterile alteration of TGMS for safeguarding seed production of twoline hybrid rice. The article also established a statistic model to conclude plant temperature by water temperatures at inflow and outflow, and air temperature and cloudage from weather station.

Development of Insect-Resistant Hybrid Rice by Introgressing the Bt Gene from Bt Rice Huahui 1 into Ⅱ-32A/B,a Widely Used Cytogenic Male Sterile System

Huahui 1 is an elite transgenic male sterile restorer line of wild rice abortive-type that expresses a Bacillus thuringiensis （Bt） δ-endotoxin and provides effective and economic control oflepidopteran insects. To exploit Huahui 1 to develop a new Bt rice, the insertion site of the Bt gene was determined by thermal asymmetric interlaced PCR （TAIL-PCR）. Bt was located in the promoter region ofLOC. Os10g10360, approximately 5.35 Mb from the telomere of the short arm of chromosome 10. For the first time, a Bt cytoplasmic male sterile （CMS） system was developed by introgressing Bt from Huahui 1. The recipient CMS system used consisted of Indonesia paddy rice-type II-32B （maintainer line） and II-32A （male sterile line）. Marker-assisted selection was used to increase selection efficiency in the backcrossing program. In BC5F1, the Bt plant 85015-8 was selected for further analyses, as it had the highest SSR marker homozygosity. In addition, the linkage drag of the foreign Bt gene in 85015-8 was minimized to 8.01-11.46 Mb. The foreign Bt gene was then delivered from 85015-8 into II-32A. The resultant Bt II-32A and Bt II-32B lines were both resistant to lepidopteran in field trials, and agronomic traits were not disturbed. The maintainability of II-32B, and the male sterility and general combining ability of II-32A, were not affected by the Bt introgression. This study demonstrates a simple and fast approach to develop Bt hybrid rice.

Genetic Mechanism of Dominant Earliness in Kefeng A,a New Rice Cytoplasmic Male Sterile Line

Kefeng A is an early maturing indica cytoplasmic male sterile （CMS） line of rice. Combinations derived from Kefeng A and late maturing indica restorer lines showed dominant earliness to various extents. To understand the genetic basis of dominant earliness, the genotype of photoperiod-sensitive genes in Kefeng A was analyzed using a complete set of heading time near isogenic lines （NILs） EGO to EG7, ER, LR, T65, T65E^b, T65E^bm, T65m, NIL（Hd1） and NIL（Hd4）. Results indicated that Kefeng A contained two dominant photoperiod-sensitive alleles E1 and Se-1^U on E1 and Se-1 loci, respectively, and the genotype of photoperiod-sensitivity genes for heading time in Kefeng A was E1E1e2e2E3E3Se-1^USe-1UEf-1Ef-1. Based on the detected heading time genotype, in combination with the heading time of Kefeng A and the early maturing phenomenon in its derived F1 hybrids, it is speculated that Kefeng A might carry a dominant inhibitor gene Su-E7 for the dominant photoperiod- sensitive gene E1, and a recessive inhibitor gene i-Se-1 for another dominant photoperiod-sensitive gene Se-1. The reason why F1 hybrids from Kefeng A exhibited early maturing was hereby analyzed and the breeding value of dominant earliness related genes in Kefeng A was discussed.

Studies of the Energy Release of Mitochondria from Sporophyte Cytoplasmic Male Sterile Rice by Differential Scanning Calorimetry

The differential scanning calorimetric (DSC)curves of the mitochondria isolated from two varieties of sporophyte cytoplasmic male sterile and their fertile lines of Yie Bai and Ma Xie type rice have been determined. The curves show that the energy is released continuously as temperature rise to 70℃. Some thermodynamic and kinetic parameters of the energy release of the mitochondria have been obtained. The presented results showed that the mitochondria from cytoplasmic male sterile rice released more heat and they had higher energy barrier, less rate, and more complicated mechanism than that of their fertile lines in the energy release process.

Influence of Genetic Drift of Restoring Gene (Rf) on Seed Purity of Yuetai A, a Honglian-Type Cytoplasmic Male Sterile Line in Hybrid Rice

The seed samples of Yuetai A, a Honglian （HL） type cytoplasmic male sterile （CMS） line in hybrid rice were investigated to assess the seed purity and to analyze the cause of off-type plants by imitating the biological contaminant to Yuetai A in Nanjing, Jiangsu Province and Lingshui, Hainan Province during 2004-2006. The seed impurity of Yuetai A mainly resulted from the genetic drift of restoring fertility gene （Rt） after biological contamination but not from its sterility unstability. All of the ten maintainer lines, five restorer lines and three thermo-sensitive genic male sterile lines used in the study could pollinate Yuetai A and Yuetai B to produce F1 plants, directly or indirectly resulting in Rf-gene drifting into Yuetai A and generating ＇iso-cytoplasm restoring-generations＇. Furthermore, high outcrossing rate and similar heading date of Yuetai A with many varieties used in rice production might easily result in the biological contamination. After removing all plants with Rf-gene mixed in Yuetai A and preventing Rf-gene drifting into Yuetai A, the seed purity of Yuetai A and Yuetai B had been raised to 100%.

Factors Influencing the Outcrossed Seed-Setting Obstacles of Photo-thermo Sensitive Genie Male Sterile Rice

Using photo-thermo sensitive genie male rice (PTGMS) Pei' ai 64S, W7415S, W6154S, N26S, Annong S, Nongken 58S, 7001S and 5088S as female parents and conventional indica lines 8258 and U89 as male parents, the factors affecting outcrossed seed-setting were analyzed. The PTGMS had obstacles in outcrossed seed setting influenced by inheritance and environment at varying degrees. Environmental temperature was regarded as the main factor that resulted in the outcrossed seed-setting obstacles. The sensitive stage was at the early stage of grain filling for outcrossed seed setting. There existed remarkable differences at the sensitivity stage, the duration of sensitive period, the sensitive level and the effective level of outcrossed seed-setting obstacles caused by environmental temperature among different PTGMS lines. Therefore, attention should be paid to outcrossed seed-setting obstacles in selection and utilization of PTGMS lines.

Analysis on Combining Ability for Characters of Male Sterile Lines in Rapeseed(Brassica napus L.)

The male sterile line is very important in the hybrid breeding program of rapeseed. This study was conducted to evaluate the combining ability of many characters of male sterile lines in Brassica napus L. Ten recessive genetic male sterile （RGMS） lines were used as parents to produce 45 single cross hybrids by using a half diallel cross method. These 45 crosses and their 10 parents were evaluated at Guiyang during 2007-2008. The results showed that both general combining ability （GCA） and specific combiing ability （SCA） effects were important for all characters, but additive gene effects were more predominant than non-additive gene effects. Qianyou 8AB and You 2894AB gave respective highly significant GCA effects of 230.94 and 127.65 kg-hm-2 for seed yield. Lines You 2894AB, QH303-4AB, You 157AB and You 2341AB gave highly significant GCA effects for oil content of 0.99, 1.62, 1.20 and 1.53%, respectively. The crosses among lines Qianyou 3AxQianyou 8B, Qianyou 8AxYou 2894B, You 2894AxQianyou 6B, Qianyou 8AxQH303-4B and Qianyou 8Ax Qianyou 6B gave high SCA effects of 616.29, 398.71,356.48, 394.24 and 303.79 kg hm-2 for seed yield, respectively. All these crosses also gave high seed yield indicating that these crosses could be used in the breeding program.

Research Progress on Fertility of Two-line Male Sterile Rice Lines

Great achievements have been made in the exploration of male sterile resources, gene mapping and cloning and molecular mechanism revealing, as well as in breeding and application of two-line rice in China. This paper briefly reviewed the discovery, types and cloning of sterile genes in rice and molecular breeding of two-line rice, and summarized the research progress of critical sterility inducing temperature, so as to provide new ideas for the research and breeding of two-line hybrid rice.

Identifying Cytoplasmic Male Sterile Types of Chinese Cabbage(Brassica campestris L.)by Molecular Markers

Different sterile cytoplasm types of nine cabbage cytoplasmic male sterile materials were identified by molecular marker in the study, in order to better use molecular marker to conduct the assisted breeding in the future. Genomic DNA was isolated from Chinese cabbage by CTAB method. The design of two pairs of specific primers was performed on conserved flanking region of orf138 gene in the GenBank. PCR was performed with genomic DNA of the nine Chinese cabbage materials. The bands were sequenced. The homologous comparison was conducted in NCBI, and finally, the type of sterile cytoplasm was determined. The results showed that the bands were amplified only in four Chinese cabbage male sterile materials with two pairs of specific primers PUPIl and PIII/PIV, while the other five materials did not obtain the relative bands. The result was consistent with the field sterility identification. And then four molecular markers of Chinese cabbage Ogura cytoplasmic male sterility （CMS） were obtained. After conducting a homologous comparative analysis with BLAST in GenBank, it was found that the homologous degree was 100% in specific segments of tbe tbree sterility materials （L1-CI, L3-CI and L3- F1 ） and Ogu orf138 gene （GenBank accession No. ： HQ149728） of the reported broccoli Ogu CMS. The homologous degree of L1-F1 was 99% with a variation point. The type of cytoplasmic male sterility of the other five materials needed further research. Four materials of the nine were identified as the radish cytoplasmic male sterility materials and four molecular markers were obtained.

Distinguishing male sterile and fertile cytoplasms in indica rice(Oryza sativa L.) by OPA 12

The majority of off-type plants in in-dica hybrid rice were CMS and itsmaintainer lines.Discrimination ofmaintainer and CMS could only beconducted at heading stage.If main-tainer and CMS line could be distin-guished earlier,it would be useful toavoid yield losses caused by usingcontaminated seeds.In this study,three pairs of A and B,Minghui 63,and Fand Fplants of Zhenshan97A/Minghui 63 were used to find