Estimation of hybrid seed production and efficient pollen flow using insect pollination of male sterile soybeans in caged plots

The aptitude of leafcutter bees to pollinate male sterile soybean plants (ms2 gene) in caged plots was evaluated in four experiments from both quantitative and qualitative points of view. The m.s. plant seed set was satisfactory: on average, it represented 60% of the male fertile isogenic seed set (with a range between 44 and 69%). The lower yield of m.s. plants was linked with a smaller number of fertile reproductive nodes. The efficient pollen flow was observed over the flowering period with both morphological and electrophoretic markers. Insect behaviour was not influenced by flower colour. Differences between flowering duration of pollen donors appear to be the major factor inducing unbalanced populations. The interest of this technique as a tool for dynamic management of the genetic variation in populations or for hybrid seed production is discussed.     

Development of thermo-sensitive cytoplasmic male sterile (TCMS) lines of wheat characterized by complete male sterility at lower-temperatures and partially restored fertility at higher-temperatures

A ‘‘two-line hybrid system’’ was developed using thermo-sensitive cytoplasmic male sterility (TCMS) from Aegilops kotschyi and is proposed as a means for seed multiplication and production of hybrid wheat. The TCMS line is crossed to a pollinator line during the normal wheat-growing season for production of hybrid wheat seeds, whereas the TCMS line is maintained through self-pollination in an environment with high temperatures. The male sterile wheat line KTP3314A, containing Ae. kotschyi cytoplasm and the short arm of chromosome 1B from Triticum spelta, was studied in different temperature regimes. KTP3314A was completely male sterile under normal wheat growing temperatures (<18 °C). Sterility of KTP3314A was partially restored when plants were grown at temperatures above 20 °C during growth stages 45–52. Male fertility was not correlated with day-length. A series of TCMS lines derived from KTP3314A showed similar environmental responses and performed well in hybrid wheat production at lower temperatures. Fertility of these TCMS lines at higher temperatures makes it possible to produce large quantities of male sterile parental seeds. Based on these findings, application of the TCMS-dependent two-line system in hybrid wheat breeding appears to have a promising future for hybrid wheat production.     

不同花粉密度条件下光温敏雄性不育小麦BS366异交结实分析

研究了人工授粉和田间不同授粉方式条件下,光温敏雄性不育小麦BS366的异交结实情况。结果表明,在不育系花后不同时间人工饱和授粉条件下,其异交结实率呈先增后减的变化,花后第4天至第8天,异交结实率达80%以上,其中第5天为88%;穗中部异交结实能力最强,但异交结实粒数变异也最为明显;多次人工饱和授粉的异交结实率达94.7%,说明该不育系具有较高的异交结实潜力;花后天数与异交结实率的关系可以用方程y=-1.457 7x2 20.489x 12.472(R2=0.914 6**)描述,观察值与模拟值相差0～9%。田间不同授粉方式下,距恢复系1.5 m内,不育系异交结实变异较大,2.0 m以外无显著差异;距恢复系0.5 m内,不育系异交结实率较高,行间差异不显著;辅助授粉时,距恢复系1.0 m以内的不育系异交结实率和杂交种子重量的增幅最大,异交结实率最高可增加45%。光温敏雄性不育小麦BS366制种时播种宽度应控制2.0 m,并在花后8 d内,尤其是花后4～8 d进行辅助授粉,可实现较高异交结实率。     

QTL mapping of male sterility-related traits in a photoperiod and temperature-sensitive genic male sterile wheat line BS366

The photoperiod/temperature-sensitive genic male sterile (P/TGMS) character is important for use of heterosis in hybrid wheat. In the present study, the fertility of 172 doubled haploid (DH) lines derived from the cross between a P/TGMS wheat line BS366 and a restorer wheat line Baiyu149 was investigated under both fertile and sterile environments during three cropping seasons. The phenotype was recorded based on the international seed setting rate. A total of 167 SSR and 1,278 SNP markers were used to construct a linkage map, with a total length of 3,748.94 cM and an average marker interval of 2.59 cM. Three QTLs were identified and designated as QF.bhw-2DS, QF.bhw-4BS and QF.bhw-7Al, explaining 6.9%–12.8%, 19.7%–25.6% and 7.2%–8.8% of the phenotypic variances, respectively. These results lay a good basis for application of male sterility-related molecular markers in improvement of two-line hybrid wheat breeding system.     

Development of dominant nuclear male-sterile lines with a blue seed marker in durum and common wheat

In order to develop genie male‐sterile lines with a blue seed marker, male‐sterile plants, controlled by a dominant nuclear gene Ms2, were used as female parents against a 4E disomic addition line ‘Xiaoyan Lanli’(2n= 44, AABBDD+4EII) as the male parent to produce monosomic addition lines with blue seed. Male‐sterile plants from the monosomic addition lines were pollinated with durum wheat for several generations and in 1989 a male‐sterile line with the blue grain gene and the male‐sterile gene Ms2 on the same additional chromosome was detected and named line 89‐2343. Using this line, the blue seed marker was successfully added to a short male‐sterile line containing Ms2 and Rht10. The segregation ratios of male sterility and seed colour as well as the chromosome figurations of different plants indicated that the blue grain genes, Ms2 and Rht10 were located on the same additional chromosome. Cytological analysis showed that the blue marker male‐sterile lines in durum wheat and common wheat were monosomic with an additional chromosome 4E. The inheritance ratio for blue seed male‐sterile plants and white seed male‐fertile plants was 19.7% and 80.3%, respectively, in common wheat. The potential for using blue marker sterile lines in population improvement and hybrid production is discussed.     

A new male sterile mutant LZ in wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Genetic male sterility (GMS) genes in wheat (Triticum aestivum L.) can be used for commercial hybrid seed production. A new wheat GMS mutant, LZ, was successfully used in the 4E-ms system for producing hybrid wheat, a new approach of producing hybrid seed based on GMS. Our objective was to analyse the genetic mechanism of male sterility and locate the GMS gene in mutant LZ to a chromosome. We firstly crossed male sterile line 257A (2n = 42) derived from mutant LZ to Chinese Spring and several other cultivars for determining the self-fertility of the F₁ hybrids and the segregation ratios of male-sterile and fertile plants in the F₂ and BC₁ generations. Secondly, we conducted nullisomic analysis by crossing male sterile plants of line 257A to 21 self-fertile nullisomic lines as male to test the F₁ fertilities and to locate the GMS gene in mutant LZ to a chromosome. Thirdly, we conducted an allelism test with Cornerstone, which has ms1c located on chromosome 4BS. All F₁s were male fertile and the segregation ratio of male-sterile: fertile plants in all BC₁ and F₂ populations fitted 1:1 and 1:3 ratios, respectively. The male sterility was stably inherited, and was not affected by environmental factors in two different locations or by the cytoplasm of wheat cultivars in four reciprocal cross combinations. The results of nullisomic analysis indicated the gene was on chromosome 4B. The allelism test showed that the mutant LZ was allelic to ms1c. We concluded that the mutant LZ has common wheat cytoplasm and carries a stably inherited monogenic recessive gene named ms1g.     

Development of photoperiod-sensitive cytoplasmic male sterile (PCMS) wheat lines showing high male sterility under long-day conditions and high seed fertility under short-day conditions

A “two-line system” using photoperiod-sensitive cytoplasmic male sterility (PCMS) caused by Aegilops crassa cytoplasm under long-day photoperiods (≧15 h) has been proposed as a means of producing hybrid varieties in common wheat (Triticum aestivum). The PCMS line is maintained by self-pollination under short-day conditions, and hybrid seeds can be produced through outcrossing of the PCMS line with a pollinator line under long-day conditions. Our previous studies revealed that PCMS lines showing complete male sterility under long-day conditions are necessary for practical hybrid wheat breeding, especially to obtain high hybrid purity in F₁ seeds. Furthermore, practical PCMS lines should have high seed fertility under short-day conditions, which is associated with female fertility. Wheat cv. Norin 26 with Ae. crassa cytoplasm exhibits high seed fertility under short-day conditions, and cv. Fujimikomugi with Ae. crassa cytoplasm shows high male sterility under long-day conditions. Here we developed practical PCMS lines derived from the F₁ generation of Norin 26 and Fujimikomugi (with Ae. crassa cytoplasm) that were then backcrossed to elite wheat lines.     

Factors responsible for levels of male sterility in photoperiod-sensitive cytoplasmic male sterile (PCMS) wheat lines

A `two-line system' using photoperiod-sensitivecytoplasmic male sterility (PCMS) caused by Aegilops crassa cytoplasm has been proposed as a newmeans of producing hybrid wheat. The PCMS line ismaintained by self-pollination under short-dayconditions (14.5 h light period), and F<sub>1</sub> seedscan be produced by outcrossing of the PCMS line witha pollinator under long-day conditions (15 h lightperiod). As the levels of male sterility in PCMSlines under the long-day conditions is a crucialfactor in determining hybrid purity of the F<sub>1</sub>seeds, a study was conducted into the effect ofseeding rate on male sterility in PCMS lines. Threedifferent density levels were tested using analloplasmic line of Japanese wheat cultivar `Norin 26'which exhibits PCMS. Levels of male sterility of thePCMS line increased at sparse planting, because tiller(ear) number per plant increased at low seedingdensity and late-appearing ears tended to exhibithigher levels of male sterility than early-appearingears. On the other hand, male sterility levels of thePCMS lines depended on genotype, e.g., the PCMS`Fujimikomugi' was completely male sterile, whereasthe PCMS `Norin 26' showed partial male sterility. APCMS line showing complete male sterility, such as thePCMS `Fujimikomugi', should produce F<sub>1</sub> seeds withhigh purity. However, the PCMS `Fujimikomugi' showeda lower female fertility. For practical use, it isnecessary to produce PCMS lines having high malesterility with high female fertility under long-dayconditions.     

棉花诱导长柱头杂交制种及配套技术研究

旨在减少棉花杂交制种用工,提高棉花杂交制种工效,降低棉花杂交种子生产成本。采用GA₃溶液诱导棉花长柱头,不去雄直接授粉;采用小型强磁高速电动机作为动力,研制小型田间快速采粉器,同时研制与采粉器配套使用的简易授粉器。通过多年的研究,研制出一种新的杂交棉制种新方法,即用100 mg/kg浓度的GA₃溶液,处理8~18天母本的幼蕾,每花点滴量150~200 μL,开花当天,不去雄父本花粉直接授予母本伸长的柱头上;研制出与之相配套的棉花快速采粉器和低温贮能式授粉器,操作简便,田间采粉速度快,授粉效率高,生产的杂交种子纯度在95%以上,符合国家良种要求。该研究将稳定的诱导技术与采粉器和授粉器结合应用于棉花杂交制种,可节省大量用工,综合提高制种工效30%以上。     

Use of herbicide-resistant genic male sterility in hybrid rice seed production

One of the difficult problems in hybrid rice seed production is the low outcrossing frequency and requirement for much labor to produce hybrid seeds. In order to simplify the process of hybrid rice seed production, herbicide-resistant photoperiod sensitive genic male sterile (HRPGMS) rice was utilized in this study. The herbicide resistance gene bar was transferred into the photoperiod sensitive genic male sterile (PGMS) rice 920S by Agrobacterium-mediated transformation and the HRPGMS line YA3530ms with good agronomic characteristics was bred by applying conventional pedigree breeding technique. The seeds of HRPGMS and pollen parent were mixed with the ratio of 4:1 in weight, and were sowed in seedling box. The mixed seedlings of HRPGMS and pollen parent grown for 30 days were transplanted by the small transplanting machine in the field. The herbicide glufosinate ammonium was sprayed at 7 days after flowering to kill all the plants of pollen parent, whereas hybrid seeds were harvested from the survived HRPGMS parent at maturity. The outcrossing frequency of HRPGMS line from two combinations in 2002 and from five combinations in 2004 were compared with a control cultivated by the conventional 2-line system. As the result, the mean outcrossing frequency in HRPGMS of the treatments were 10.6–24.5% compared with 5.5% in PGMS of the control in 2002, and that were 24.7–32.0% compared with 7.5% in the control in 2004. Consequently, using HRPGMS in two-line system was proved to be a new method that would simplify the process of hybrid rice seed production and to increase outcrossing frequency without any artificial supplementary pollination processes.     

Characterization of a male sterile mutant from progeny of a transgenic plant containing a leaf senescence-inhibition gene in wheat

A male sterile plant of wheat (Triticum aestivum L.) segregated from progenies of a transgenic family containing the leaf senescence-inhibition gene P _SAG12 -IPT in the genetic background of ??Xinong 1376??, a well adapted winter wheat cultivar. The male sterile plant (named TR1376A) showed no phenotypic changes, except for florets and male organs, compared to its male fertile sibling plants (named TR1376B). The glumes and florets of male sterile TR1376A plants widely opened whereas those of the fertile counterpart TR1376B were closed or opened only briefly at flowing. Anthers of TR1376A were slender and indehiscent, and failed to release pollen. Compared to TR1376B, TR1376A anthers contained greatly reduced amounts of pollen, which was inviable or weakly viable. Ultra-structure studies indicated that cells in the endothecium and middle layers of the anther wall were dissolved or poorly developed in the sterile anthers of TR1376A. Molecular studies showed that the male sterility of TR1376A was caused by a sequence deletion or mutation that occurred in the promoter region of the transgene. F₁ hybrids of TR1376A and TR1376B gave 1:1 segregation of male fertility to sterility, indicating that the male sterility of TR1376A was heritable and controlled by a single dominant gene (named Ms1376). To date, only a few dominant nuclear male sterility genes have been characterized and one of them (Ms2) has been successfully used to improve wheat cultivars through recurrent breeding strategies. The discovery of the Ms1376 gene provides another dominant male sterile source for establishing recurrent breeding systems in wheat.     

Hybrid seed set in wheat is a complex trait but can be improved indirectly by selection for male floral traits

Efficient hybrid wheat breeding requires the redesign of the wheat floral architecure to enhance cross-pollination. Several studies evaluated the phenotypic variation and the genetic architecture of male floral traits, but their contribution to the most important trait, hybrid seed set on the female parent, has not yet been considered. To bridge this gap, we employed 31 male lines and evaluated the hybrid seed set on two female tester lines in crossing blocks. Hybrid seed set showed large genetic variance and high heritability, which demonstrates the potential for the improvement of this trait. However, the assessment of hybrid seed set is difficult as secondary traits like plant height and especially flowering time, as well as the environment largely influence the hybrid seed set. Nevertheless, a moderately high correlation between visual anther extrusion and hybrid seed set opens up the possibility to use visual anther extrusion as an indirect trait for preliminary male screenings. Further research evaluating traits influencing female receptivity coupled with genomics-assisted approaches are highly recommended to develop an improved selection portfolio for maximizing hybrid seed set.     

水稻小粒不育系新组合卓两优141混播制种优势分析

传统的杂交水稻制种方式落后、效益低、成本高的问题,制约了杂交稻推广应用。利用小粒型不育系可实现杂交水稻混播、混收、机械分离等全程机械化制种,降低杂交稻制种成本,提高制种效率。本研究以小粒型不育系卓201S、大粒恢复系R141和新组合卓两优141为材料,对其适合机械化制种的农艺性状、异交特性、混播制种及大面积制种实践进行了研究。结果表明,与C815S相比,卓201S株高较矮,穗长较长且直立,包颈粒率、穗萌率、黑粉病、开颖率均低于C815S,对"九二〇"较敏感,且较难落粒,具有较好的农艺性状。而R141株高较高,花粉数量大,花期长,且对环境温度较钝感,两者均具有良好的异交特性。卓201S谷粒厚1.71mm,千粒重14.00g,R141谷粒厚2.23 mm,千粒重28.20 g,两者籽粒大小差异显著,通过特定的狭长形筛孔筛子,实现了杂种F1与父本种子的高效分离,杂交种子含父本率为0,杂种损失率为2.31%,种子纯度在生产上达标,可实现机械化制种;与传统制种模式相比,混播制种可使父本基本苗减少85%,母本容量增加20%,制种产量增加21.37%,制种综合效益增加31.4%。父母本较好的农艺性状...     

Cytological and genetical studies of a male sterile celery

Summary The inheritance and nature fo male sterility of a cerley (Apium graveolens L.) strain (MS1) is reported. Male sterility in MS1 is determined by a a recessive genotype for a single locus. Tests for linkage with the isozyme chromosome markers SDH-1 and PGM-1 were negative. MS1 male sterility was associated with a defective tapetum characterized by prominent vacuoles and premature degeneration. The stamens in the male sterile strain persisted in the flowers up to stigma receptivity, while in the male fertile plants they dropped before style expansion. The male sterile flowers produced normal amounts of nectar, resulting in cross pollination by various species of pollinators. It was estimated that the sterile strain produced 30% less seed than normal male fertile. Its possible use for hybrid celery seed production is discussed.Research supported by grants from the California Celery Research Board and BARD I-483-82.     

Some aspects of cross-pollination in wheat (Triticum aestivum L.) 1. Pollen concentration in the field as influenced by variety,diurnal pattern,weather conditions and level as compared to the height of the pollen donor

Summary For the large-scale production of hybrid seed the cross-fertilizing capacity of wheat must be increased. Breeding good pollinators is, therefore, a necessity. Five spring wheat varieties were examined for differences in pollen shedding capacity under normal field conditions. The examination was carried out by collecting pollen on object slides every hour from 06.00 till 20.00 h during flowering. In general, the variety Peko proved to be the best shedder of pollen, while Opal was the worst. Gaby, Jufy I and Orca were intermediate. Consistent results were hard to obtain mainly due to varying weather conditions.It was established that pollen is released throughout the whole period of daylight. The maximum release of pollen, however, occurred from 08.00 till 11.00 h. A less pronounced and much smaller peak was noticed from about 15.00 till 18.00 h. Most pollen appeared to be released at a temperature of 16–20°C and a relative humidity of 70–75%. Although there seems to be some relationship between the diurnal pattern of the pollen release and the indicated values of these meteorological factors, one cannot say that the former is entirely determined by the latter.It has been suggested in the literature that a male-sterile parent that is shorter than its pollinator would be most likely to produce good seed set in the hybrid wheat seed production field. Investigations showed that a greater concentration of pollen was found below the ear level of pollinator plants than at or above ear level.     

绿豆雄性不育突变体msm2015-1的遗传学与细胞学分析

单产水平低一直是我国绿豆存在的产业问题。绿豆杂种优势显著是提高产量的有效途径,而雄性不育材料是实现杂种利用的宝贵资源。本研究利用~(60)Co-γ辐射诱变技术首次在绿豆中获得雄性不育突变体,并命名为‘不育绿豆2015-1’(male sterile mungbean2015-1,简写为msm2015-1)。msm2015-1营养生长期表型与‘苏绿1号’没有明显差异。花器官形态正常,但花药发白且不能正常开裂散粉,成熟期座荚率几乎为0。遗传分析显示,育性分离群体中可育与不育分离比符合3∶1,表明不育性状由单个隐性核基因控制。花粉发育的细胞学分析发现, msm2015-1的盛开花朵中花粉粒几乎未在柱头附着,体外亦未萌发。花粉细胞质未被Alexander染液正常着色, I_2-KI染色出现典败、圆败和染败3种类型。DAPI染色显示, msm2015-1花粉细胞核发育异常。醋酸洋红染色表明,败育发生在花粉发育早期,花粉母细胞在四分体时期减数分裂的不对称和异常多分裂是导致花粉败育的主要原因。     

Natural male sterile mutant in hot chilli (Capsicum annuum L.)

Summary A natural male sterile mutant was recovered from the normal population of chilli cv. CA 452-1. The male sterility in the mutant was governed by a single recessive gene ms. The excessive vegetative growth provided easy identification of male sterile plants in the seedling stage. The plant was found to be promising for hybrid seed production.     

Electron-microscopy on anther tissue and pollen of male sterile and fertile wheat (Triticum aestivum L.)

Summary 1. Anther tissue and pollen grains of cytoplasmic male sterile and fertile wheat were studied with the aid of an electron-microscope for differences in cytoplasmic structures.2. Clear and consistently occurring cytoplasmic differences were not observed, although unidentifiable particles were discovered in both the sterile and the fertile material.3. Male sterility finds expression in the small number of organelles in the sterile pollen grains which consequently are little or hardly physiologically active and thus subject to a rather rapid degeneration. On the contrary the fertile pollen grains are full of cell organelles and therefore well equipped for a complete development.4. The fertile pollen grain contains a clear intine which is thicker than the exine. Only a very thin intine was observed in the sterile material.5. Male sterility has no visible influence on exine formation. Ubisch bodies occur in the anthers of both male sterile and fertile plants. There is no sign of any contribution to exine formation on the part of the Ubisch bodies. There was more evidence of exine formation from within than from without. The normality of exine formation of sterile pollen grains need not indicate exine formation from without; it suggests rather that exine is largely formed before degeneration.     

甘蓝型隐性核不育杂交油菜制种中父本和可育株花粉授精竞争力的研究

用花时指示性状作父本，隐性核不育系1601A（圆叶）作母本，父母本行比为1：10，在花期不拔除可育株让其父本和母本中可有株花粉对其不育株自由授粉，其父本花蕾总数占该群体可育株花蕾总数的17.3%的情况下，父本花粉授精竞争能力较强，不育株所结种子花叶株率达34.94%，父本花粉授精竞争力为1.02。其中，靠近父本的不育株所结种子花叶株率达80%，竞争力达3.63。母本可育花蕾虽占总花蕾的82.7%，但不育株所结种子中圆叶株率略只占65%，其母本中的可育株花粉授精竞争力为-0.27,而且即使母本中的可育株所结的种子，在靠迫父本的数行中，其父本花粉的竞争力仍为正值，强子可育株本身花粉的竞争力。     

Reproductive barriers in the intergeneric hybridization between <Emphasis Type="Italic">Chrysanthemum grandiflorum</Emphasis> (Ramat.) Kitam. and <Emphasis Type="Italic">Ajania przewalskii</Emphasis> Poljak. (Asteraceae)

In order to reveal the causes of low seed set in the intergeneric cross between Chrysanthemum grandiflorum (Ramat.) Kitam. ‘Zhongshanjingui’ (female parent) and Ajania przewalskii Poljak. (male parent), we systematically investigated megasporogenesis and megagametogenesis of female parents, pollen germinability of male parents, pollen–pistil interaction and hybrid embryo development using paraffin section and light, fluorescence and scanning electron microscopy. The results indicate that development of megagametophyte of female parent belongs to Polygonum type and 84.6% ovules developed to normal 8-nucleate embryo sac at the stage of pollination; the pollen germinability in vitro of male parent was 39.3%; a large number of pollen grains germinated normally and no callose deposited on the female stigmas within 24 h after pollination. Furthermore, about 75% ovules were fertilized, whereas only 17.2% ovules developed to normal maturing embryos at 15 days after pollination due to abortion at various developmental stages. Taken together, these results suggest that sterile ovules of female parent, pollen viability and pollen–pistil interaction have no significant influence on seed set in the cross. A high percentage of embryo abortion or degeneration, i.e. post-fertilization barrier, may be the main factor for low seed yield.