Nitrogen Fixation by Hybrids of White Clover (Trifolium repens L.) and Trifolium nigrescens

Development of hybrids between white clover ( Trifolium repens L.) and Trifolium nigrescens provides a novel route for genetically improving the reproductive capacity of white clover, provided the hybrids are agronomically viable, particularly with respect to N₂ fixation. A comparative study of growth and rates of N₂ fixation over 21 days was conducted with the parental species, F ₁ hybrids and backcross hybrids, in flowing solution culture, without a supply of mineral N to the plants. T. nigrescens was unable to fix N₂ in association with the strains of Rhizobium leguminosarum biovar. trifolii selected for inoculation. Rates of N₂ fixation per plant increased in the order T. nigrescens < F ₁ hybrid < T. repens < backcross 1. Specific rates of N₂ fixation (days 0–21) increased in the order T. nigrescens < F ₁ hybrid < backcross 1 <  T. repens . Dry matter production and nodule biomass per plant increased at a higher rate in backcross 1 hybrids than in T. repens. The results suggest that the potential for N₂ fixation by backcross 1 hybrids is at least as great as that by T. repens .     

Growth and reproductive characteristics in backcross hybrids derived from Trifolium repens L. × T. nigrescens Viv. interspecific crosses

Interspecific hybridisation with the close relative, Trifolium nigrescens (Ball clover) is a possible strategy to achieve increased reproductive potential of white clover (Trifolium repens). Fertile F₁ plants have been used as the basis for two generations of backcrossing to T. repens as the recurrent parent. F₁ and backcrossed plants were assessed in both glasshouse and field for a range of morphological traits, including inflorescence production, and the level of water soluble carbohydrates in the stolons. Plants resulting from two generations of backcrossing had an increased allocation of dry matter to inflorescence production in comparison with T. repens. Variation within these plants for agronomic traits (e.g. stolon length, dry weights etc.) suggests that selection for these traits is feasible and in combination with increased inflorescence production offers a potentially valuable approach to germplasm improvement in white clover. This revised version was published online in August 2006 with corrections to the Cover Date.     

Morphological characteristics of hybrids between white clover, Trifolium repens L., and Caucasian clover, Trifolium ambiguum M. Bieb

Hybrids between the stoloniferous Trifolium repens (2n = 4x = 32) and the rhizomatous Trifolium ambiguum (2n = 4x = 32) have been produced using T. repens as the recurrent parent. Morphological characteristics of the parent species and the F₁, BC1, BC2 and BC3 hybrids were examined in a glasshouse experiment. Leaflet ratios and general plant shape indicated that the BC3 hybrid was similar to T. repens. Separation of plants into components of above- and below-ground growth showed that T. repens had a greater total plant dry weight than T. ambiguum but 24% of its dry weight was in roots compared with 22% in roots and 44% in rhizomes in T. ambiguum. The BC3 generation contained plants that were predominantly T. repens-like with stolons, but also had a small proportion of their total dry weight (3%) as rhizome, confirming the potential to produce plants that combine stoloniferous and rhizomatous growth. The proportions of rhizome and stolon varied within the BC3 generation, enabling farther selection and plant breeding. The BC3 hybrids produced similar numbers of inflorescences to T. repens however, fertility was lower, although there was some variation within the BC3 for both characters.     

Wide hybridization between wheat (Triticum L.) and lymegrass (Leymus Hochst.)

A total of 240 F₁ hybrids was made beween wheat (Triticum aestivum L. em. Thell. (2n = 6x = 42) and T. carthlicum Nevski (2n = 4x = 28)) and perennial lymegrass (North European Leymus arenarius (L.) Hochst. (2n = 8x = 56) and North American L. mollis (Trin.) Pilger (2n = 4x = 28)). The wide crosses yielded embryos in 20% of caryopses and 96% of the embryos developed into normal hybrid plants. The hybrids were vegetatively vigorous, with evidence of the Leymus rhizomatous habit. Those deriving from L. arenarius survived overwintering in Iceland, but the hybrids L. mollis did not, whereas in a milder environment, both showed perenniality. Cytogenetic analysis of root tip cells before the plants were treated with colchicine showed that 21 out of 28 hybrids investigated had chromosome mosaics, with a population of both amphihaploid and amphidiploid cells. This spontaneous doubling of somatic chromosomes occurred in all cross combinations, with the highest average frequency of diploid cells (28%) in T. carthlicum × L. arenarius crosses. A few selfed seeds have been obtained from a T. aestivum × L. arenarius hybrid. All the hybrids were treated twice with colchicine, but the treatment appeared to have little or no effect on the frequency of chromosome doubling in the hybrids deriving from T. aestivum. The frequency of diploid cells, however, increased significantly (e.g. to 80%) in the hybrids deriving from the T. carthlicum parent. Genomic in situ hybridization confirmed the hybridity of the plants and showed that the hybrids were amphiploids containing genomes of both wheat and lymegrass. In situ hybridization using ribosomal DNA probe differentiated chromosomes of L. mollis, L. arenarius from those of wheat. The hybrids are being backcrossed with lymegrass pollen, aiming to domesticate the wild, perennial species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Meiosis and fertility in interspecific hybrids of Nicotiana tabacum L. ×N. glauca Grah. and their derivatives

Nicotiana glauca is of potential interest to breeders as it carries resistance to black root rot of tobacco. Cytological investigations of sexual interspecific hybrids of N. tabacum T′T′TT (2n = 4x = 48) cv. ‘Wiślica’ × N. glauca GG (2n = 2x = 24) were carried out. The analyses of chromosome association at diakinesis and metaphase I in the PMCs of amphihaploid F₁ T′TG (2n = 3x = 36) revealed low variable pairing with 0–5 bivalents. The sterile amphihaploids F₁ were converted into partial female fertile amphidiploids T′T′TTGG by chromosome doubling. Among 36 mature plants obtained, 15 were found to have chromosome numbers (2n = 6x = 72) and were verified as amphidiploids, 9 had (2n = 6x = 70 or 71) chromosomes while the remaining 12 were haploid. True amphidiploids, in spite of quite high chromosome pairing during meiosis, were very different in pollen fertility, ranging from 0% to 85%. Male fertility disturbances did not correlate with the degree of female fertility upon pollination with N. tabacum. Sesquidiploids T′TG (2n = 5x = 60) obtained from backcrossing the amphidiploids to parental tobacco showed more than 22 bivalents, 10–12 univalents and occasional multivalents that indicated the possibility of interchange events between N. tabacum and N. glauca genomes.     

普通小麦与野生大麦的属间杂交

通过活体/离体幼胚培养获得了智利大麦(Hordeum chilense,2n=2x=14)、海大麦(H.marinum,2n=2x=14)及平展大麦(H.depressum,2n=4x=28)与普通小麦(Triticum aestivum,2n=6x=42)之间的属间杂种。智利大麦×小麦及海大麦×小麦杂种在形态上偏向父本小麦,而平展大麦×小麦杂种除穗部性状外,其形态明显偏向母本。所有杂种均自交不孕     

小麦属2个种间杂种及3个亲本的核型分析

【研究目的】本研究旨在通过细胞学鉴定判断小麦属2个种间杂种的真实性,了解其亲本的染色体组及倍数性;【方法】本试验采用了常规染色体组型分析方法;【结果】分析和报道了小麦属2个种间杂种及其3个亲本的核型,根据核型分析的有关参数,阐明了两个杂种1894×Ps5、新7×Ps5及其亲本新7、1894和Ps5的核型特征,其核型分别为：新7(2n=6x=42)：2M+12sm+28m(2SAT)(2B),1894(2n=6x=42)：2M+20sm+18m(2SAT)+2st(2B),Ps5(2n=4x=28)：2M+12sm(2SAT)+12m+2st(2B),1894×Ps5(2n=5x=35)：2M+19m+12sm+2st(2B),新7×Ps5(2n=5x=35)：1M+21m+13sm(2B);【结论】由此鉴定了2个杂种的真实性--均为真杂种,并确认了亲本新7和1894的第一套和第二套染色体均分别为A组和B组,倍数性均为6x,它们的第三套染色体属什么染色体组有待进一步研究。     

Introgression as a Route to Improved Drought Tolerance in White Clover (Trifolium repens L.)

Backcross hybrids between the important forage legume white clover Trifolium repens , which is stoloniferous, and the related rhizomatous species T. ambiguum have been produced using T. repens as the recurrent parent. The drought tolerance of parents and two generations of backcross plants, possessing both stolons and rhizomes, was studied in deep soil bins in a glasshouse both as monocultures and in mixtures with perennial ryegrass. Soil moisture content, leaf relative water content, and leaf water potential were measured on plants subjected to drought cycles of 4 weeks duration and those watered normally and maintained at field capacity. Six cycles of drought, carried out over 2 years, showed that T. ambiguum and the first and second generation backcross hybrids maintained a greater leaf relative water content and higher leaf water potential than T. repens at comparable levels of soil moisture. The dry matter yield of T. repens was significantly reduced over each drought cycle compared to the watered treatments, while the yield of T. ambiguum increased over the 4-week cycle in both treatments. The yield of the backcross generations was intermediate between those of the two parental species. The implications of these results for white clover germplasm improvement programmes are discussed.     

Analysis of stability for some characters in Kabuli Chickpea

Summary The meiotic behaviour of a hybrid between Triticum aestivum and the amphiploid Hordeum chilense x T. turgidum conv. durum, was studied using a C-banding staining method. This hybrid has the genome formula of AA BB D Hch with 2n=6x=42 chromosomes. The durum wheat chromosomes (genomes A and B) were easily recognized, whereas the D and Hch chromosomes were recognized as a whole. Meiotic pairing was homologous, as expected (14 bivalents from A and B genomes +14 univalents from D and Hch genomes). However, some pollen mother cells at metaphase-I presented pseudobivalents that could have been caused by either homoeologous or autosyndetic pairing amongst D and Hch chromosomes.     

钙离子胁迫对3种牧草种子萌发及幼苗生长的影响

以白车轴草、紫云英、紫花苜蓿种子为材料,通过测定发芽率、发芽势、发芽指数、恢复萌发率等指标,研究钙离子浓度对3种牧草种子萌发及幼苗生长的影响,以期为喀斯特地区牧草栽培提供科学参考。研究结果表明:1)低浓度钙离子胁迫(0～0.09 mol·L^-1)对3种牧草种子发芽影响有限,而高浓度钙离子胁迫(0.12～0.15 mol·L^-1)能够明显抑制3种牧草种子萌发,且低浓度钙离子(0.03 mol·L^-1)胁迫可以促进紫花苜蓿种子萌发; 2)白车轴草、紫云英和紫花苜蓿种子发芽指数均随钙离子浓度增加逐渐减小; 3) 3种牧草种子的恢复萌发率表现为紫花苜蓿>紫云英>白车轴草; 4)钙离子浓度明显抑制3种牧草幼苗的生长,低浓度钙离子(0.03 mol·L^-1)有利于紫花苜蓿幼苗生长。可见,紫花苜蓿对高钙环境的适应能力要好于白车轴草和紫云英,是喀斯特地区较为适宜的栽培牧草品种。     

Incompatibility in diploid and tetraploid crosses of Cucumis sativus and Cucumis metuliferus

The African horned cucumber (Cucumis metuliferus Naud.; 2x = 2n = 24) contains genes that can confer resistance to many important cucumber (C. sativus L.; 2x = 2n = 14) pests [e.g., root-knotnematode, Meloidogyne incognita (Kofoid & White) Chitwood]. Cucumber is highly susceptible to this root-knot nematode species, and a recent screening of C. sativus accessions in the U.S. National Plant Germplasm collection did not identify sources of resistance. Thus,autotetraploids of Cucumis sativus and C. metuliferus were created to recover fertile resistant interspecific progeny. Autotetraploids were obtained at the highest rate when seeds were immersed in 0.5% colchicine for a period of 6 to 8 hrs. Treatment durations less than 6 hrs produced few tetraploids, and durations of 10 hrs or more were lethal to seeds or developing seedlings. Crosses between C. sativus and C. metuliferus were made using diploid and tetraploid lines in all possible combinations, including reciprocals. Fruit development occurred in crosses when diploid and tetraploid C. sativus were used as the female parent. However, seeds developed only in fruit of C. sativus (4n) ×C. metuliferus (2n) crossings. Seeds from these crosses, however,were flat and not viable. No fruit development occurred in crosses whereC. metuliferus was used as the female parent. This revised version was published online in August 2006 with corrections to the Cover Date.     

Production of 27-, 28-, and 56-chromosome apomictic hybrid derivatives between pearl millet (2n=14) and Pennisetum squamulatum (2n=54)

Summary An interspecific hybridization program designed to transfer gene(s) controlling apomixis from Pennisetum squamulatum Fresen. (2n=6x=54) to induced tetraploid (2n=4x=28) cultivated pearl millet, Pennisetum americanum (L.) Leeke resulted in four offtype plants, two with 27 chromosomes and two with 28 chromosomes. These plants were found among 217 spaced plants established from open-pollinated seed of an apomictic 21-chromosome polyhaploid (2n=21) plant derived from an apomictic interspecific hybrid (2n=41) between tetraploid pearl millet and Pennisetum squamulatum. It appeared that a 21- (or 20-) chromosome unreduced egg from the apomictic polyhaploid united with a 7-chromosome pearl millet (2n=2x=14) gamete to produce a 28- (or 27-) chromosome offspring. Meiotic chromosome behavior was irregular averaging from 3.60 to 4.05 bivalents per microsporocyte in the 27- and 28-chromosome hybrids. The 27- or 28-chromosome hybrids, like the 21-chromosome female parent, shed no pollen, but set from 1.8 to 28 seed per panicle when allowed to outcross with pearl millet. Progeny of the 28-chromosome hybrids were uniform and identical to their respective female parents, indicating that apomixis had been effectively transferred through the egg. In addition, a 56-chromosome plant resulting from chromosome doubling of a 28-chromosome hybrid was identified. Pollen was 68 per cent stainable and the plant averaged 2.3 selfed seeds per panicle. Chromosomes of the 56-chromosome plant paired as bivalents (x=10.67) or associated in multivalents. Three to nine chromosomes remained unpaired at metaphase I. Multiple four-nucleate embryo sacs indicated the 56-chromosome hybrid was an obligate apomict. The production of 27-, 28-, and 56-chromosome hybrid derivatives were the results of interspecific hybridization, haploidization, fertilization of unreduced apomictic eggs, and spontaneous chromosome doubling. These mechanisms resulted in new unique genome combinations between x=7 and x=9 Pennisetum species.     

重金属胁迫对白三叶种子萌发的影响

为了探讨重金属胁迫对白三叶(Trifolium repens)种子萌发的影响,本研究模拟4种不同浓度重金属离子(Zn²⁺、Pb²⁺、Cu²⁺、Cd²⁺)条件,测定种子的发芽率、发芽势、发芽指数和相对胚根(芽)长等相关指标。结果表明:白三叶种子对Cd²⁺的耐受能力最强,Zn²⁺和Cu²⁺次之,对Pb²⁺的耐受能力最弱。总体上,种子的发芽率、发芽势、发芽指数和相对胚根(芽)长随着重金属离子浓度的增加表现出不同程度的下降趋势。高浓度重金属离子对白三叶种子萌发及生长发育会产生严重的阻碍作用。然而,低浓度的Zn²⁺、Cu²⁺、Pb²⁺对胚的生长具有一定促进作用,表明白三叶对低浓度重金属环境有一定适应性。在本试验中,白三叶对Cd具有较强的耐受能力,5种不同浓度的Cd²⁺条件下种子的发芽率均高于50%,且Cd²⁺浓度为1mg/L时可促进种子的萌发。此外,进行过重金属处理的种子会表现出相对胚根长小于相对胚芽长,表明重金属胁迫首先影响种子根部的生长发育,从而抑制白三叶的生长。综上所述,白三叶对低浓度的Zn²⁺、Pb²⁺、Cu²⁺、Cd²⁺和高浓度Cd²⁺具有较好的耐受能力,可作为上述环境的绿化植被,并且在重金属Cd土壤污染中有良好修复前景。     

Assessment of genetic diversity in clover species from Sardinia, Italy, using AFLP analysis

Two species, Trifolium glomeratum and T. nigrescens, from Sardinia, Italy, were analysed for genetic diversity using amplified fragment length polymorphism (AFLP). Variation between and within populations was compared between the inbreeder, T. glomeratum, and the outbreeder, T. nigrescens. Four AFLP primer combinations resulted in a total of 292 loci, of which 75% were polymorphic in T. glomeratum and 85% in Trifolium nigrescens. Variation was highest between populations in both species, but the difference between populations was greater in T. glomeratum (Fst = 0.17), compared with T. nigrescens (Fst = 0.02). Cluster analysis and principal coordinates analysis were used to verify the relationships found. The high level of genetic variation within populations in both species is attributed to the movement of sheep between paddocks, the existence of both species in Sardinia for thousands of years and the persistence of a long‐lived seedbank due to the production of large numbers of small seeds with high levels of hard seededness.     

基于SSR标记的贵州野生白三叶遗传多样性分析

采用SSR分子标记技术,对贵州22份野生白三叶(Trifolium repens)样品遗传多样性及亲缘关系进行了研究.结果表明,利用20对SSR引物共扩增出202条带,其中169条带具有多态性,多态性位点达83％;根据SSR标记的多态性计算不同采样地野生白三叶样品之间的遗传距离(GD),22份样品之间遗传距离介于0.140 6～0.630 4之间,说明样品间遗传差异较大;聚类分析结果,当相似系数为0.64时,22份白三叶被分成3类:第1类只有W14,表明W14与其余材料的亲缘关系较远;第2类包括W2、W4、W8、W18居群,表明这4个居群相似性较大,遗传距离较近;第3类包括17个居群,居群间有着复杂的遗传关系.研究结果表明,22份贵州野生白三叶的遗传多样性十分丰富.     

Cytogenetics of a new trispecies hybrid in cotton: [(Gossypium hirsutum L. × G. thurberi Tod.)2 × G. longicalyx Hutch. & Lee]

A three‐species hybrid named HTL including Gossypium hirsutum L. [2n = 4 x = 52, (AD)₁ genome] was created using the pseudophyletic introgression method with G. longicalyx Hutch. & Lee (2n = 2x = 26, F₁ genome) as donor parent and G. thurberi Tod. (2n = 2x = 26, D₁ genome) as bridge species. The new hybrid was totally self‐sterile and its interspecific status was confirmed using simple sequence repeat markers and cytogenetic analysis. Cytogenetic studies showed that its chromosome configuration was 2n = 52 = 14.13 I + 15.10 II + 1.03 III + 0.9 IV + 0.03 V + 0.13 VI (where I, II, III, IV, V and VI are univalents, bivalents, trivalents, tetravalents, pentavalents and hexavalents, respectively). Prospects for successfully exploiting the HTL hybrid in breeding programmes are discussed.     

低温处理对白三叶叶绿素、可溶性糖和丙二醛等的影响

为探索低温胁迫对白三叶的叶绿素、可溶性糖和丙二醛等生理指标的影响,为白三叶的抗寒性育种和资源的合理利用提供基础数据,试验以新品系08-2和海法白三叶(Trifolium repens L.‘haifa’)为材料,依次设置10℃、5℃、0℃、-5℃、-10℃、-15℃等6个处理,研究低温下叶绿素、可溶性糖和丙二醛等的变化,对材料的抗寒性进行评价。结果表明,变温对白三叶的叶绿素、可溶性糖和丙二醛等的影响较大。随着温度的降低,白三叶的叶绿素含量总体呈现降低的趋势,在较低(≤0℃)温度处理下,08-2的叶绿素含量降低速度小于海法的;08-2的可溶性糖含量在0℃时达到最高,海法的在5℃达到最高;丙二醛含量和电导率逐渐升高,但08-2变化幅度低于海法。综合分析表明,新品系08-2耐寒性优于海法白三叶。     

Chromosome doubling and fertility study of Alstroemeria aurea × A. caryophyllaea

Self-pollinations of a diploid (2n = 2x = 16) interspecific hybrid Alstroemeria aurea × A. caryophyllaea, resulted in no seeds. Backcrosses of the hybrid with parent A. aurea did not produce any seeds. In an attempt to restore the hybrid fertility, an efficient in vitro procedure has been developed and applied effectively in the chromosome doubling of the diploid hybrid. Forty-one percent of the treated plants were proven to be truly tetraploid by chromosome counts and stomatal measurements after applying 0.2 to 0.6% colchicine for 6 to 24 hours. Over 87.5% of these colchicine-induced tetraploids were stable and retained their tetraploidy after one year of growth. The fertility of the hybrid was not restored although the pollen stainability was increased from 0 to 12% after chromosome doubling. Cytological studies on the pollen mother cells (PMCs) of the sterile diploid hybrids revealed abnormal meiotic behaviors. In addition, aneuploid chromosome numbers, ranging from 2n = 1 to 2n = 18, were observed in over 45% of the PMCs examined. PMCs of the colchicine-induced tetraploids showed that meiotic chromosome pairings were normal in most cases (1.59_I + 15.07_II + 0.05_III + 0.03IV). These results indicate that the sterility of this hybrid is not only caused by parental chromosome differences, but other complex fertility/sterility-regulating mechanisms are involved too. This revised version was published online in July 2006 with corrections to the Cover Date.     

Development of high-gossypol cotton plants with low-gossypol seeds using trispecies bridge crosses and in vitro culture of seed embryos

The objective of this work is to develop an upland cotton, Gossypium hirsutum L. [2n = 4x =52, 2(AD)h], having a reduced level of gossypol in the seeds for food and feed uses, and a high level of gossypol in the remaining organs to limit pest incidence. Using G. sturtianum Willis (2n = 2x = 26, 2C1) as donor and G. thurberi Torado (2n = 2x = 26, 2D1) or G. raimondii Ulbrich (2n = 2x = 26, 2D5) as bridge species, two trispecies hybrids G. thurberi– G. sturtianum– G. hirsutum and G. hirsutum– G. raimondii– G. sturtianum were synthesized. Both trispecies hybrids were male sterile. Recurrent backcrossing to G. hirsutum as pollinator and selfing of the second backcross (BC) progenies resulted in seeds which were rescued by in vitro culture. In total, 1208 flowers of the trispecies hybrids and their BC progenies yielded 192 seed embryos from which 62 plants were obtained. Cytogenetic analyses indicated a relatively high frequency of chromosome pairing and chiasmata. The gland levels in backcross seeds ranged from glandless seeds to normally glanded seeds. All vegetative parts of those hybrids were glanded, but a wide range of variability for gland density was observed on leaf, stem, bract and calyx. Plants derived from seeds having a reduced level of gossypol constitute very interesting germplasm to develop a cultivated glanded cotton with low-gossypol seeds. This revised version was published online in July 2006 with corrections to the Cover Date.     

长穗偃麦草—普通小麦核质杂种的创制及其细胞遗传学分析

以长穗偃麦草(Elytrigia elongata=Agropyron elongatum,2n=70)为母本,普通小麦为父本,进行核置换回交,现已获得第14次回交的核质杂种材料。根据育性和生活力的表现以及细胞遗传学分析,长穗偃麦草-普通小麦核质杂种可分为以下类型:(1)生长正常,育性较好,染色体构型为21″W+1′Eev;(2)生长正常,完全不育或育性很低,染色体构型