Identification and characterization of important sterile and maintainer lines from various genotypes for advanced breeding programmes of onion (Allium cepa)

Onion is one of the major vegetable crops in terms of production as well as consumption. In the current research, available onion genetic stock was evaluated to identify male-sterile lines and produce high-yielding F₁ hybrids for future breeding programmes. A mitochondrial DNA-based marker was mapped and correlated with phenotypic traits to isolate male-sterile plants. Based on the floral and pollen structure, nine putative male-sterile lines were identified. On the other hand, for nuclear marker identification at Ms locus, two sets of primers were used, one for Ms dominant allele and another for sterile and maintainer plants. Results revealed that 70% of open pollinated varieties (OPVs) possess plants with sterile cytoplasm coupled with genetic sterility at Ms locus, called sterile “A” line. Approximately 20% of plants in some genotypes were identified with normal (N) cytoplasm having recessive fertility gene at Ms locus, called maintainer “B” line. Based on the present findings, “A”, “B” and “R” (restorer line), future F₁ hybrid seed production systems in onion is discussed.     

大葱成熟胚愈伤组织诱导及植株再生

为了建立大葱组培高频再生体系,以‘铁杆大葱’的成熟胚为材料,探讨培养基种类、植物生长调节剂浓度等因素对大葱成熟胚愈伤组织诱导及再生的影响。结果表明,MS培养基适合大葱成熟胚愈伤组织诱导,大葱成熟胚愈伤组织诱导最适宜的培养基为MS+2,4-D (2~3 mg/L)。大葱成熟胚愈伤组织分化培养最适培养基为MS+6-BA (1.0 mg/L)+KT (1.0 mg/L)+NAA (0.1 mg/L),分化率达90%;对分化的再生苗移植于1/2 MS培养基上进行生根及壮苗培养,生根率达到100%,炼苗移栽后小苗的成活率达到90%以上。本研究为进一步优化大葱再生体系提供了依据,为后续大葱的遗传转化的研究起了推动作用。     

甘蓝型油菜白花突变体的遗传研究和白花胞质雄性不育系的选育

用白花突变体和白花不育突变体分别选育白花品系G95150和白花胞质不育系7600 A。遗传研究表明,白花对黄花由一对不完全显性基因控制。7600A在农艺性状上与其保持系G95150完全一致：叶色浓绿,植株较矮（163.5 cm）、生育期较长（243天）、抗寒能力较差（冻害率100%,冻害指数33.8）。7600A花色纯白、花瓣皱缩、花药小、低温存在微量花粉。G95150和7600 A的芥酸含量为0.8854 %,硫甙含量70.5461μmol/g。     

洋葱细胞质雄性不育系及保持系小孢子发生的细胞学观察

为揭示洋葱细胞质雄性不育系小孢子的败育时期,利用石蜡切片技术对洋葱细胞质雄性不育系8A及其保持系8B的小孢子发育过程进行观察和比较。结果表明：石蜡切片洋葱细胞质雄性不育系8A和保持系8B造孢组织时期小孢子的生长发育没有明显的差异;洋葱不育系8A花药绒毡层在花粉母细胞时期就与中层完全分离,并逐渐膨大挤压花粉母细胞,最后逐渐降解。其降解后的残余物侵入药室,与小孢子混合粘连在一起,占据药室的一部分空间,并且不能供给小孢子生长发育所需要的营养物质,最终小孢子降解,花药败育。试验明确了洋葱细胞质雄性不育系8A的小孢子败育时期为花粉母细胞时期。因此,在洋葱雄性不育系8A的不育机理研究中,应重点在花粉母细胞时期进行深入研究。     

Limitations of PCR‐based molecular markers to identify male‐sterile and maintainer plants from Indian onion (Allium cepa L.) populations

The main aim of this study was to validate PCR markers for determining cytoplasm and genotypes at the Ms locus in short‐day onion. Three cytoplasmic (OSN, MKFR and accD) and four nuclear (OPT, jnurf13, AcSKP1 and AcPMS1) markers were employed. Sel. 121‐1 had 100% S cytoplasm, whereas Sel. 121‐2, ‘Pusa Red’ and ‘Pusa Madhavi’ had 88%, 33% and 17% S cytoplasm, respectively. ‘Early Grano’ and ‘Pusa Riddhi’ did not possess S cytoplasm. Observations in 33 commercial varieties revealed two with sterile (S) cytoplasm. Nuclear markers were not found in linkage disequilibrium with the Ms locus, and the constitution of Ms alleles by OPT was different from other three markers, which were in conformity with each other. The other three markers predicted that most of the plants should be homozygous recessive. Anther colour also did not confirm the sterility status. It can be concluded that accD may be used for cytoplasm determination based on the ease of its use. For the Ms locus tagging, more markers are needed to be evaluated to isolate maintainer lines from open‐pollinated populations.     

A male sterile line with dominant gene (Ms) in cabbage (Brassica oleracea var. capitata) and its utilization for hybrid seed production

A male sterile plant, 79-399-3, was identified from a spring cabbage line 79-399 in 1979. Light-microscopic studies with paraffin section indicated that meiotic division stopped at the tetrad stage. The ratio of male sterile plants to fertile plants in the progenies of test crosses was 1:1 in five experiments conducted in 1982 and 1991–1994. However, some male sterile plants were sensitive and developed a very low number of viable pollen grains. When sensitive male sterile plants were selfed, the progenies segregated into male sterile and fertile plants at a ratio of 3:1. The male sterile plants from selfing of sensitive male sterile plants were propagated by tissue culture and crossed with different inbred lines. From the progenies of the crosses, populations with 100% male sterile plants were observed. The results indicated that homozygous dominant male sterile (MsMs) plants segregated from the selfed progenies. Populations with 100% male sterile plants and stable male sterility were developed as male sterile lines. Several favorable combinations were also selected by crossing the male sterile lines with inbred lines with high combining ability for desirable horticultural characters. This revised version was published online in July 2006 with corrections to the Cover Date.     

HL-CMS同质异核近等基因恢复系‘L-Rf6’的构建

为了研究红莲型水稻中2个恢复基因与2个不育基因的互作关系,用红莲型水稻不育系‘粤泰A’(‘YTA’)与恢复系‘9311’杂交,所得F1代再与保持系‘粤泰B’(‘YTB’)多次回交,最终构建了与不育系‘YTA’同质异核的近等基因恢复系‘L-Rf6’。根据Rf5和Rf6的基因序列设计了特异性分子标记,并对‘L-Rf6’进行检测。结果显示成功构建了近等基因恢复系‘L-Rf6’。对近等基因恢复系的不育基因orfh79的表达量进行分析,结果显示‘L-Rf6’中orfh79的表达量与‘YTA’比较明显下降。该近等基因恢复系的构建为阐述红莲型水稻多个恢复基因与多个不育基因的互作关系打下基础。     

Mitochondrial genome variation in Allium ampeloprasum and its wild relatives

Limitation in mitochondrial genome diversity of leek, revealed by restriction fragment length polymorphism (RFLP) analyses with mitochondrial gene probes, prevent a cytoplasmic male sterility (CMS) system in elite populations. However, mitochondrial genome diversity was detected in Allium ampeloprasum L. wild accession and landraces, as well as in pearl onion. Within this plant material, nine mitotypes were distinguished and could be used in order to broaden the genetic basis of leek. A chimeric mitochondrial gene configuration is usable as a marker for the sterility inducing cytoplasms (S₁) in chives (Allium schoenoprasum L.) and in onion (Allium cepa L.) for (S) and (T) cytoplasm. This chimeric mitochondrial gene configuration is also present in the subgenus Allium, revealed by polymerase chain reaction (PCR). However, only a faint amplicon was observed in a few accessions investigated herein, suggesting that this fragment might be present to a lesser level in mitochondrial DNA, as a sublimon.     

洋葱雄性不育恢复位点Ms座位的SSR标记开发及其应用

本研究旨在开发稳定可靠的洋葱育性位点分子标记,并将其应用于育种实践,筛选育种系,从而缩短育种周期,节省育种成本,加速洋葱杂交种的培育进程。以洋葱育性恢复Ms座位的AFLP序列为基础,采用序列比对分析、PCR检测和表型分析等方法,开发、鉴定、应用了WH-SSR-1分子标记。结果表明:WH-SSR-1标记与Ms位点紧密连锁,Ms位点基因型为纯合显性MsMs时,有1条102 bp的扩增带;纯合隐性msms时,有1条99 bp的扩增带;杂合Msms时,有102 bp和99 bp两条扩增带。32份洋葱材料的验证结果证实了Ms座位的标记类型与基因型完全相符。随后从4个OP群体中直接获得了2个配套的不育系与保持系,‘吊玉’和‘天正红玉’因未检测到保持株或不育株,无法简单实现配套。WH-SSR-1标记与Ms位点紧密连锁,能够将其用于育种系筛选的育种实践,从OP群体中直接选择保持株和不育株,同时实现两系配套。     

Cytoplasmically controlled cold tolerance in a cytoplasmic-genetic male sterile line of rice

Summary The cytoplasmic-genetic male sterile line, Krishna-A with Kalinga-I cytoplasm was developed in rice through repeated backcrossing the completely pollen sterile F₁ of the cross Kalinga-I/Krishna with the recurrent male parent cv. Krishna. The germination percentage and root-shoot length in Krishna-A at 12°C and 9°C indicated a higher degree of tolerance to low temperature than the cold tolerant female parent Kalinga-I. The high yielding cold tolerant variety Kalinga-I was developed from a cross involving the cold tolerant variety Dunghansali and the high yielding variety IR 8. It is inferred that tolerance to low temperature during germination in Krishna-A was inherited from the cytoplasm of cv. Dunghansali through Kalinga-I besides male sterility. The cytoplasmic control of cold tolerance is reported here for the first time in rice literature. The male sterile line Krishna-A because of its tolerance to low temperature would be suitable for the development of hybrid rice especially for areas where low temperature is a problem during germination.     

大葱生长发育过程中可溶性糖、可溶性蛋白质及游离氨基酸含量变化规律的研究

研究了大葱生长发育过程中可溶性糖、可溶性蛋白、游离氨基酸含量变化动态。结果显示,随着大葱的生长,可溶性糖呈现低-高-低-高的变化规律,但不同材料的变化有差异。日本品种假茎可溶性糖含量变化幅度不大。而地方品种胭脂红、托县孤葱在采收时假茎中可溶性糖含量较高。假茎蛋白质含量表现为高-低-低-高的趋势。定植30 d后假茎蛋白质含量最高,以后随着生长、发育,蛋白质含量逐步下降;到采收期假茎内蛋白质含量又在积累,葱叶中蛋白质含量变化与假茎相似。大部分材料定植后随着大葱的生长发育,假茎中游离氨基酸含量呈上升趋势,近采收期达到最大值。不同材料葱叶中游离氨基酸含量不同,随着生长发育呈现低-高-低-高的变化趋势,葱叶中游离氨基酸含量远远低于假茎。     

Conversion of the genic male sterility (GMS) system of bell pepper (Capsicum annuum L.) to cytoplasmic male sterility (CMS)

Non‐pungent bell pepper (Capsicum annuum L.) lacks the cytoplasmic male sterility (CMS) nuclear restorer allele, Rf, and CMS cannot be employed in its F₁ hybrid seed production. To demonstrate that the genic male sterility (GMS) system in non‐pungent bell pepper can be converted to the CMS male sterility system, the conversion of GMS to CMS for non‐pungent bell pepper line GC3 was conducted by introgression of S‐type cytoplasm and the Rf allele from tropical pungent donors. After morphological traits were evaluated, two lines from BC₁F₁ containing S‐type cytoplasm and four lines from BC₂F₂ containing Rf allele, phenotypically similar to GC3, were obtained and could be employed as CMS male sterile lines and restorer lines for non‐pungent bell pepper. Four molecular markers potentially linked to traits of interest were also evaluated in BC₁F₁ and BC₁F₂ populations. This is the first time that GMS has been successfully converted to CMS in bell pepper, a significant contribution for bell pepper hybrid seed production.     

Morphological and cytological study in a new type of cytoplasmic male-sterile line CMS-GIG2 in sunflower (Helianthus annuus)

Cytoplasmic male sterility (CMS) is essential for sunflower hybrid production. CMS-GIG2, a new sunflower CMS type, was further confirmed by crossing with the maintainer and restorer lines for the CMS-PET1, both of which maintain the male sterility of CMS-GIG2. Meiotic division in CMS-GIG2 was observed with 4'6-diamidino-2-phenylindole dihydrochloride staining, indicating that microspore formation was disrupted before the meiotic cytokinesis was completed. Light microscopy observation showed that both middle layer and tapetal cells expand radially rather than degrade over time, followed by failure to form tetrads and normal microspores. This morphological defect leading to male sterility in CMS-GIG2 differs from that observed in the PET1 CMS type. CMS-GIG2 will certainly provide additional genetic diversity for sunflower hybrid breeding programmes.     

Characterization of different cytoplasmic male sterility systems in Indian mustard (Brassica juncea L. Czern & Coss)

Six cytoplasmic male sterility (CMS) systems, viz. moricandia, ogura, oxyrrhina, siifolia, tournefortii and trachystoma of Indian mustard (Brassica juncea L.) were characterized for agronomic and floral characteristics. Introgression of alien cytoplasm caused alterations in different floral traits in ogura, siifolia, tournefortii and trachystoma CMS systems. Varied response to different genetic backgrounds of CMS lines indicated the presence of cytoplasmic–nuclear interaction in alteration of floral traits. On the basis of floral characteristics, CMS systems could be grouped into distinct classes. Siifolia, tournefortii and trachystoma CMS lines had narrow petals, while moricandia, ogura and oxyrrhina had wider petals, which were distinguishable on the basis of visual observations. The ratio between length and width of petals were >2.0 in wide petal group but <2.0 in narrow petal group. Further, the relative position of anther and stigma, which was estimated as the ratio between stamen and style length could differentiate the CMS systems. Stamens were longer than styles in oxyrrhina, equal in moricandia and shorter in ogura, siifolia, tournefortii and trachystoma male sterile lines. Non‐viable pollen grains were present in moricandia and oxyrrhina systems, but absent in other systems. In tournefortii and trachystoma, few flowers showed petaloid corolla. All male sterile lines, except trachystoma, which showed crooked siliqua formation were at par with their respective maintainers for flower initiation, plant height, primary branches, seeds per siliqua, seed yield, harvest index, oil and protein content. In general, flower senescence and maturity occurred earlier in male sterile lines than in their respective maintainer lines.     

Fertility alteration in the photo-thermo-sensitive male sterile line BS20 of wheat (Triticum aestivum L.)

The photo-thermo-sensitive male sterile (PTMS) wheat line BS20 showed male sterility under short-day photoperiod and low temperature conditions. Photo-thermo-controlled growth chambers were used to study the fertility alteration characteristics of PTMS wheat line BS20. The period of photo-sensitivity and thermo-sensitivity was coincident with the pollen development stages from pollen mother cell (PMC) formation to mono-nucleate. Fertility alteration of BS20 was induced mainly by temperature, and the critical point of temperature was 10–12^∘C. Below the critical temperature, BS20 remained sterile and was not affected by photoperiod. However, above the critical temperature, the fertility of BS20 improved as the temperature rose. In the temperature range of 12–14^∘C the fertility of BS20 improved significantly as the photoperiod extended.     

Genetics of fertility restoration in A2-based cytoplasmic genetic male sterility system in pigeonpea (Cajanus cajan)

The three short duration cytoplasmic genetic male sterility (CGMS) hybrids developed using A₂ (Cajanus scarabeoides) cytoplasm-based male sterile lines (CORG 990047A, CORG 990052A and CORG 7A) and the restorer inbred AK 261322 and their segregating populations (F₂ and BC₁F₁) were subjected to the study of inheritance of fertility restoration in pigeonpea. The fertility restoration was studied based on three different criteria, namely, anther colour, pollen grain fertility and pollen grain morphology and staining. The F₂ and BC₁F₁ populations of the three crosses, namely, CORG 990047A × AK 261322, CORG 990052A × AK 261322 and CORG 7A × AK 261322, segregated in the ratio of 3:1 and 1:1, for anther colour (yellow:pale yellow), pollen grain fertility (fertile:sterile) and for pollen grain morphology and staining. The above study confirmed that the trait fertility restoration was controlled by single dominant gene. This finding can be utilized for the identification of potential restorers, which can be further used in the development of CGMS-based hybrids in pigeonpea.