Cytology and reproductive behavior of diploid, tetraploid and hexaploid germplasm accessions of a wild forage grass: Paspalum compressifolium

Summary Fourteen germplasm accessions of Paspalum compressifolium native from southern Brazil were cytologically and embryologically analysed. The study revealed that one accession was diploid (2n=20), twelve were tetraploid (2n=40) and one was hexaploid (2n=60). This is the first report of diploid and hexaploid cytotypes for this species. Studies on microsporogenesis, megasporogenesis, and embryo sac development indicated that the diploid cytotype had regular meiotic behavior and reproduces sexually. Tetraploid cytotype usually had an important proportion of chromosomes that associated as quadrivalents during meiosis and reproduced by mean of aposporous apomixis. The hexaploid cytotype showed irregular meiotic behavior with about one third of the chromosomes associated as multivalents and reproduced by aposporous apomixis. Thus, P. compressifolium could be an agamic complex. Breeding in this complex is possible due to the presence of diploid sexuals which can be treated by colchicine and pollinated by apomicts.     

Chromosome numbers and microsporogenesis in Brachiaria brizantha (Gramineae)

The genus Brachiaria, native to the African tropical savannas, has achieved significance as a pasture grass in many tropical and subtropical countries, including Brazil. Many species and accessions are polyploid and apomictic, which complicates the improvement of breeding stocks through hybridization. In support of breeding programs, cytogenetic characterization, including chromosome counts and evaluation of the meiotic behavior in the accessions of the Brachiaria has been undertaken at the Embrapa Beef Cattle Center. In this study, 22 accessions of B. brizantha were analyzed of which one was found to be diploid (2n = 2x = 18), 18 were tetraploid (2n = 4x = 36) and three were hexaploid (2n = 6x = 54). The meiotic chromosome behavior was slightly irregular in the diploid and in some tetraploid accessions, and highly irregular in most tetra- and hexaploid accessions. Meiotic abnormalities were those common to polyploidy, i.e., multivalent chromosome association at diakinesis and irregular chromosome segregation leading to micronuclei formation in the tetrad stage. Low frequencies of multivalent chromosome associations among polyploids suggest that they may be segmental allopolyploids. This revised version was published online in August 2006 with corrections to the Cover Date.     

Failure of cytokinesis and 2n gamete formation in Brazilian accessions of Paspalum

Cytological analysis on 112 Brazilian accessions of Paspalum showed that 52 were affected by failure of first or second cytokinesis during microsporogenesis. When the failure of cytokinesis occurred only in the first division and in the second division the cytokinesis was normal, a dyad of microspores was formed. In most of them the two cells remained binucleate, but in some a restitutional nucleus was observed. When the first division was normal and the failure of cytokinesis occurred only in the second division, dyads and triads were observed. A restitutional nucleus in one or both cells of the dyad, or in the binucleate cell of the triad was also observed. The percentage of dyads and triads varied among accessions, reaching 15.75 of the sporads in some. The absence of cytokinesis indicates the possibility that the widespread occurrence of polyploidy in the genus Paspalum originates from 2n pollen grains. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic diversity for agronomic and fiber traits in day-neutral accessions derived from primitive cotton germplasm

Upland cotton (Gossypium hirsutumL) is an important crop that is cultivated in warm climates through-out the world. Agronomic performance and fiber quality must continually be improved if cotton is to maintain economic viability. Primitive ancestors of cotton contain diversity for trait improvement; however, many of these accessions have a short-day flowering response (photoperiodic) and are not readily useable in breeding programs. In this study, 114 day-neutral derived primitive germplasm lines were evaluated in field trials for two years. Agronomic and fiber trait data were collected and analyzed. Variance components, genotypic values, and genotypic correlations were calculated. Genotypic effects for all traits studied made significant contributions to the phenotypic variation indicating genetic diversity among these lines. The predicted genotypic values showed a wide range of variation for agronomic and fiber traits. Weak genotypic correlations were found between yield and 2.5% span length and fiber strength, two important fiber traits. Although these day-neutral derived accessions had lower lint percentage, they had improved fiber length, strength, micronaire, and comparable yields with two commercial cultivars. Thus, these day-neutral derived accessions are sources of genetic variation that when used in breeding programs offer the potential to improve important traits and expand genetic diversity.Contribution of the USDA-ARS in cooperation with the Mississippi Agric. and Forestry Exp. Stn. Mention of trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by USDA, ARS and does not imply its approval to the exclusion of other products or vendors that may also be suitable.     

Chromosome number and microsporogenesis in a pentaploid accession of Brachiaria brizantha (Gramineae)

Chromosome number, microsporogenesis and mode of reproduction are described for an accession of Brachiaria brizantha, a grass of African origin. Cytological analysis revealed that the accession BRA005886 is pentaploid (2n = 5x = 45), with a base number of x = 9. Multivalent chromosome associations, from tri‐ to pentavalents, were recorded in diakinesis, and the further meiotic behaviour was highly irregular as well. Most abnormalities were related to irregular chromosome segregation commonly found in polyploids. Micronuclei were observed following telophases I and II. Some micronuclei near the cell wall were released as microcytes, whereas others remained in the tetrad. Other meiotic abnormalities, such as cell fusion and the absence of cytokinesis causing the formation of dyads and triads were also recorded. Binucleate microspores and 2n microspores resulting from nucleus restitution were observed among the normal ones. Limitations of this accession for use in hybridization programmes are discussed.     

Evaluation of Paspalum germplasm for variation in leaf wax and heat tolerance

Summary The genus Paspalum contains a number of important forage grasses, including P. dilatatum Poir. (dallisgrass) and P. notatum Flugge (bahiagrass). Even though many Paspalum species are adapted to moist, humid areas, they frequently are subjected to extended periods of drought stress which reduces forage productivity. This study was initiated to determine the range of variation for different indicators of drought tolerance in several Paspalum species. Fifteen species representing a broad cross section of germplasm were grown in a replicated field nursery and sampled for leaf epicuticular wax content on four dates over three years. Wax was extracted with chloroform and quantified by the dichromate oxidation technique. Heat tolerance (solute leakage technique) was also determined by calculating the ratio of the conductivity of the external bathing solution of leaves following a heat treatment to the conductivity of the bathing solution after leaves were boiled. Wax values of the entries ranged from 5.7 to 0.88 mg/dm². Both relative rankings and absolute wax values were very consistent across all sampling dates. The accessions of Paspalum nicorae Parodi and P. quandrifarum Lam. were consistently high in wax content, while the accessions of P. commune Lillo and P. unispicatum (Scribn. et Merr.) Nash were consistently low. The heat tolerance testing demonstrated that the accession of P. notatum var. saurae Parodi was the most heat tolerant of the entries. Results from this investigation demonstrate the wide range of diversity that exists for both epicuticular wax and heat tolerance within the genus Paspalum.     

Mating biology,nuclear DNA content and genetic diversity in spider plant (Cleome gynandra) germplasm from various African countries

Cleome gynandra (L.) Briq. is an African leafy vegetable with a potential to improve food security and micronutrient deficiencies. Cytological traits, breeding biology and genetic diversity of 30 selected entries of C. gynandra from six African countries were investigated. The entries consisted of advanced lines, gene bank accessions and farmers’ cultivars. Our study revealed chromosome numbers of 2n = 34 in root tip metaphase cells from one entry. The 30 entries were found to be diploid with genome sizes ranging from 2.31 to 2.45 pg/2C. Hand pollination experiments were carried out to assess self‐incompatibility within the entries and revealed that they are self‐ and cross‐compatible. For genetic diversity studies within and among the entries, the pooled data of 499 polymorphic bands from 11 amplified fragment length polymorphism primer combinations and nine simple sequence repeat markers were used. The genetic distance among the entries ranged from 0.13 to 0.77. In a principal coordinate analysis, the farmers’ cultivars formed a cluster separate from the advanced lines and the gene bank entries, and the latter were not well resolved.     

Direct as well as indirect somatic embryogenesis from immature (unemerged) inflorescence of a minor millet Paspalum scrobiculatum L.

Somatic embryos differentiated directly on the rachis of immature inflorescences of Paspalum scrobiculatum L. cv. PSC 1 on culture to MS or N₆ medium supplemented with different concentrations (4.5–22.5 μM) of 2,4-dichlorophenoxyacetic acid (2,4-D). Direct embryogenesis on the rachis of inflorescence explants forms the first instance in graminaceous plants. Highest frequency of direct embryogenesis (34%and 30% cultures, respectively) was possible on N₆ medium supplemented with 4.5 μM of 2,4-D and MS medium fortified with9.0 μM of 2,4-D. Other tissues of the explant, floral-primordia, only after an initial phase of callusing differentiated into somatic embryos; indirect embryogenesis. Somatic embryogenesis, direct as well as indirect, was resolved by scanning electron microscopy. The somatic embryos germinated and developed into plantlets on regeneration medium. Interestingly, one week incubation of somatic embryos on activated charcoal (0.5%) fortified basal medium, supported high potential for ‘germination’ on transfer to charcoal-free basal medium. This beneficial effect of activated charcoal on regeneration of somatic embryos into plantlets is the first record in the Gramineae. This revised version was published online in July 2006 with corrections to the Cover Date.     

Isolation of two microsatellite markers from BAC clones of the Vf scab resistance region and molecular characterization of scab‐resistant accessions in Malus germplasm*

A polymerase chain reaction (PCR)‐based method was developed to isolate microsatellite markers from large‐insert genomic DNA clones of bacterial artificial chromosome (BAC) libraries. The method is fast and economic since it does not require subcloning. It was applied to isolate a microsatellite marker from a BAC clone of the chromosomal region containing the apple scab resistance gene Vf. The Vf gene of Malus floribunda 821 is the most widely used source of scab resistance in apple breeding. A second microsatellite was found on the extremity of a BAC clone flanking the Vf locus. The two microsatellites allowed the identification of the presence of the Vf gene in the scab‐resistant accessions M. micromalus SA573‐3, ‘Golden Gem’, M. prunifolia 19651 and MA 16 not previously known to carry this gene. They were also used to verify the correctness of the published genealogical tree of the Vf cultivar ‘Florina’, in which a probable mistake was identified. This analysis shows the importance of genotyping the Vf locus when choosing scab‐resistant germplasm as parents in breeding programmes.     

Cytogeography and variation of stomatal size of Paspalum Glaucescens (Gramineae; Paniceae) in Southern Brazil

Paspalum glaucescens belongs to the informal group Plicatula, reproductively characterized by the dominance of tetraploid apomitic lines in most of its populations, with rare diploid, sexual counterparts. The species shows high phenotypic variation. Twenty nine Southern Brazilian accessions were cytologically and morphologically analysed. Most of the accessions were tetraploid (2n = 4x = 40). Meiotic study of three tetraploids showed their irregular behaviour. Eight accessions presented the diploid level (2n = 2x = 20).This can be considered a very high frequency of diploids in a member of the Plicatula group. While the tetraploids are concentrated at the highest elevations, diploids were detected mostly in the lowlands. Average stomatal sizes were quite distinct on different ploidy levels, being larger in tetraploids. This revised version was published online in July 2006 with corrections to the Cover Date.     

燕麦不同品种染色体倍性的检测

研究了6个不同品种燕麦的染色体倍性差异,通过根尖染色体计数法和气孔保卫细胞横纵径及保卫细胞叶绿体计数法等方法对燕麦染色体倍性进行了观察鉴定.结果表明,6个不同品种燕麦中定燕1号、晋燕8号、晋燕17号、宁莜1号为六倍体(2 n=6 x=42),白燕6号、白燕7号为四倍体(2 n=4 x=28),同倍性不同种之间气孔保卫细胞叶绿体数目相近,不同倍性间气孔保卫细胞的大小差异较明显.经X2适合性检测,六倍体和四倍体植株气孔保卫细胞横径均值之比和保卫细胞叶绿体数的平均值之比符合其染色体数目之比3∶2,而纵径均值比值不符合.     

Development of colchicine‐induced tetraploid St. Augustinegrass (Stenotaphrum secundatum) lines

St. Augustinegrass is well suited for lawns and commercial landscapes. While many genotypes are cross‐fertile, all cultivars are propagated vegetatively in sod production. To ensure varietal purity, development of sterile triploid hybrids by crossing tetraploid and diploid genotypes has been successfully used in other warm‐season turfgrasses. Applying this model in St. Augustinegrass would be beneficial to sod producers and turf managers who require purity for certification and uniformity for performance, respectively. This study was conducted to develop colchicine‐induced tetraploid lines of St. Augustinegrass. Seeds of cultivar ‘Raleigh’ were treated with four colchicine concentrations at four exposure times. A non‐treated control was included among the treatments. Seedlings that germinated were screened for genome size changes using flow cytometry. Line DSA 13005 and two progeny lines derived through selfing, DSA 16001 and DSA 16016, were corroborated as tetraploids (2n = 4x = 36) through chromosome counts. These lines will be used in future breeding efforts to attempt development of sterile triploid cultivars of St. Augustinegrass.     

Chromosome doubling of androgenic haploid plantlets of rice (Oryza sativa) using antimitotic compounds

The regeneration of haploid plantlets is considered as a bottleneck in rice anther culture. In this study, an antimitotic chromosome doubling method, simple and efficient, of androgenic haploid plantlets resulted in an efficient doubled haploid obtainment. Through chromosome doubling capacity comparison of the three antimitotic compounds (colchicine, trifluralin and oryzalin), colchicine at 500 and 625 mg/L without supplementing with DMSO was found to be the best antimitotic treatment, with a chromosome doubling capacity of 40%. Furthermore, the in vitro growth of plantlets was followed to analyse the effects of antimitotic compounds. Colchicine treatments were more toxic than dinitroanilines, and colchicine DMSO-supplemented treatments had significant lower values on shoot growth. On the other hand, dinitroaniline compounds impeded root growth, provoked helical growth of shoot and caused the apparition of white nodules in the base of the plantlet due to sprouting abortion. In this study, a protocol for doubled haploid plant recovery was established taking advantage from androgenic haploid plantlets in order to increase the number of doubled haploid plantlets produced after an anther culture protocol.     

ZZWW型四倍体雌蚕减数分裂中的性染色体行为

用过冷却方法处理得到的ZZWW型四倍体雌蚕与ZZ型二倍体交配 ,根据亲、子代个体蚕的标记性状表现及性染色体数观察 ,探讨在减数分裂中ZZWW雌蚕的性染色体的联会、分配的行为方式和概率大小 ,推测ZZWW型四倍体雌蚕在减数分裂中的性染色体联会、分离的可能方式。拟为多倍体诱发机理的解析、交配后代表现的预测提供理论依据和为多倍体研究实用化开发提供有效途径。     

Identification of the trisomic series in diploid Solanum tuberosum L., group tuberosum. I. Chromosome identification

Summary The morphology of 12 pachytene bivalents of the dihaploid Solanum tuberosum cv. Gineke is described, and the morphological features which are helpful for a critical chromosome identification are illustrated. The difficulties of identifying the individual chromosomes and the extra chromosomes in trisomics are discussed.     

Assessing genetic diversity in the cassava (Manihot esculenta Crantz) germplasm collection in Brazil using PCR-based markers

Knowledge of the origin, organization and nature of the cassava (Manihot esculenta Crantz) germplasm collection in Brazil is incomplete due to lack of critical information on several aspects of the collection. This study verifies the utility of SSR-primed PCR markers for germplasm assessment and then utilizes these markers as well as RAPD's to characterize the Brazilian collection. We specifically address the following questions: 1)what is the relationship of morphologically closely related species to cultivated cassava? 2) What is the genetic diversity of cultivars within and between different habitats in Brazil? 3) Do agronomic traits and molecular markers reveal the same relationship among cassava accessions? 4) How complete is the Brazilian cassava collection and how well is it represented in the Word Core Collection of cassava, maintained by CIAT? Results of the interspecific studies of cassava and its wild relatives confirms the close relationship of cassava, Manihot esculenta ssp. esculenta to Manihot esculenta ssp. flabellifolia as well identifying several other closely related wild species. Next, PCR-based markers indicate a strong grouping of varieties related to the region of cultivation in Brazil. Moreover, important regions such as Cerrados and Amazon are relatively poorly represented in germplasm collections. Interestingly, the relationships of accessions based on agronomic traits are not fully congruent with relationships revealed with RAPD markers. Finally, the genetic diversity of the Brazilian cassava collection is not fully represented in the Core of the Word Core Collection of CIAT.     

The physiology of variation in the time of ear emergence among wheat varieties from different regions of the world

Summary Differences in response to photoperiod and vernalization and genetic variation independent of photoperiod and vernalization (earliness per se), affecting time of ear emergence of wheat, were identified in controlled environment experiments with 33 varieties of diverse geographical origin. The results were compared with an analysis of time of ear emergence of 10409 T. aestivum accessions from the USDA Small Grain Collection grown from autumn sowings in Pendleton, Oregon, and spring sowings in Fargo, North Dakota. The effect of differences in photoperiod and vernalization sensitivity on time of ear emergence was similar to the effect of earliness per se, both under controlled environment conditions and in the field. Most of the accessions from low latitude regions reached ear emergence rapidly owing to their insensitivity to photoperiod and vernalization and earliness per se factors accelerating ear emergence. Lateness was common among accessions from Northern Europe, Afghanistan and Turkey, which was due to sensitivity to photoperiod and vernalization, and to earliness per se factors delaying ear emergence. The physiological basis of earliness per se is discussed.     

Genetic characterization and identification of new accessions from Syria in an olive germplasm bank by means of RAPD markers

Thirty-two olive cultivar accessions from Syria, most of them obtained from collecting expeditions, were characterized by means of RAPD markers before being introduced in the World Germplasm Bank of Cordoba. A total of 79 polymorphic bands(6.1 polymorphisms per primer) out of 93(7.1 bands per primer) were scored for the13 primers used, corresponding to 84.9% of the amplification products. Thirty-one different genotypes were clearly discriminated. Differences were not found among the amplification profiles from different individuals of the same cultivar. Only two cases of mislabeling or errors of planting were found. Fourteen accessions corresponding to 6 homonyms were discriminated by RAPDs as different genotypes. The dendrogram obtained by RAPD analysis included three major groups. Some evidence of relationships of the Syrian accessions studied according to their geographic origin and/or diffusion was found. For instance, cultivars from the Central Syria (Tadmur/Palmyra)such as Toffahi', ‘Abbadi Abo Gabra’-1033,‘Abo Kanani’, ‘Shami’-1041, ‘Abbadi Shalal’ ‘Adgam’-844 and ‘Majhol’-1013 clustered in Group 1 and 2. Six cultivars from Northern Syria clustered in Group 2. But it was not found a geographic structure for the cultivars from South and West of Syria. These results agree with the hypothesis of autochthonous origin of most of the olive cultivars. Some associations between cultivars from Central Syria and their fruit size were observed. This suggests that fruit size was a criterion of local selection in olive cultivars of this area. This revised version was published online in July 2006 with corrections to the Cover Date.     

Evidences for the control of chromosome number variation by a programmed-cell-death-like pathway in citrus callus

Chromosome number variations were found in citrus callus induced from diploid genotypes. Further cytological examination of single-protoplast-derived cell lines showed that these chromosome number variations arose spontaneously from mitotic irregularities, but that the ploidy level remained relatively stable over one year in culture. The frequent occurrence of tetraploid cells and the stable maintenance of ploidy level seemed paradoxical. To explain this, the mitotic indexes of diploid and tetraploid sub-populations were calculated, respectively. The results showed that the division of tetraploid cells was inhibited. The results of a TUNEL (TdT-mediated dUTP Nick-End Labeling) assay, DNA staining and comet-electrophoresis further suggested that tetraploid cells were eliminated by a programmed-cell-death-like pathway. An erratum to this article is availbale at .     

Chromosome location and microsatellite markers linked to a powdery mildew resistance gene in wheat line 'Lankao 90(6)'

Wheat lines known as 'Lankao 90(6)', derived from the cross 'Mzalenod Beer' (hexaploid triticale)/'Baofeng 7228'//'90 Xuanxi', carry a recessive powdery mildew resistance gene temporarily named PmLK906 . Gene PmLK906 appears to be different from known wheat powdery mildew resistance genes. PmLK906 was tagged using microsatellite markers in a segregating population derived from the cross 'Chinese Spring'/'Lankao 90(6)21-12'. The dominant microsatellite marker Xgwm265-2AL was linked in repulsion with PmLK906 at a genetic distance of 3.72 cM, whereas the co-dominant Xgdm93-2AL was linked to PmLK906 at a genetic distance of 6.15 cM. Both markers were placed on chromosome arm 2AL using 'Chinese Spring' nulli-tetrasomic lines. The recessive PmLK906 has a different specificity to the dominant resistance alleles located at the Pm4 locus and appeared to be located to a locus different from Pm4 .