Identification and genetic diversity analysis of date palm (Phoenix dactylifera L.) varieties from Morocco using RAPD markers

Genetic variation among 43 date palm (Phoenix dactylifera L.) accessions, including 37 accessions from Morocco and 6 cultivars from Iraq and Tunisia, was studied using Random Amplified Polymorphic DNA (RAPD) markers. The pre-screening of 123 primers on four genotypes allowed selection of 19 primers which revealed polymorphism and gave reproducible results. All 43 analysed genotypes were distinguishable by their band patterns. RAPD technology therefore appears very effective for identifying accessions of date palm. RAPD-based genetic distance was used to determine the relationships between the accessions. The grouping-association identified by cluster analysis was rather weak. However, morphologically similar varieties clustered together. A relatively low polymorphism and a lack of evident organisation are observed among the date palm varieties grown in Morocco. This could be related to the mode of introduction and maintenance of the Moroccan date palm germplasm involving limited foundation germplasm, exchange of cultivars between plantations, and periodic development of new recombinant cultivars following sexual reproduction. This revised version was published online in August 2006 with corrections to the Cover Date.     

Increasing the rate of ripening of date palm fruit (Phoenix dactylifera L.) cv. Helali by preharvest and postharvest treatments

‘Helali’ is a late season date palm cultivar. At the mature (Bisir) stage, the fruit are astringent as a result of high contents of soluble tannins, and removal of tannins is necessary for the fruit to be edible. During the harvesting season, only 30–40% of the total fruit might normally ripen (Rutab stage) on the tree and the remaining fruit fail to ripen. This study showed that bunch bagging with different materials such as black or blue polyethylene bags, white ‘agrlsafe’ (polypropylene fleece) and paper bags during the growing season significantly increased the rate of fruit ripening and increased Rutab yield per bunch. In this respect, black and blue polyethylene bags were the most effective followed by ‘agrlsafe’ and paper bags. Preharvest ethrel application significantly increased Rutab fruit yield per bunch compared to the controls. There were no significant differences in Rutab yield per bunch between sprays or injection of ethrel into the bunch peduncle. Postharvest dipping of fruit at the Bisir stage in ethrel at 4.2 ml/l and abscisic acid at 1.0 mM significantly enhanced ripening, compared to the controls. However, ABG-3168 (an ethylene blocker) application at 3.33 g/l significantly inhibited ripening, suggesting a role for ethylene in the ripening process. Ethanol vapor significantly hastened ripening of Bisir fruit over 10 days at ambient conditions in desiccators. The response of immature fruit (according to fruit density and TSS) to ethanol vapor was much greater than mature ones. Also, immersion of fruit in water for 10 h significantly increased fruit ripening compared to the controls, but to a lesser extent. It is concluded that ‘Helali’ date ripening could be hastened by bunch bagging during growth, or by exposing the Bisir fruit to ethanol vapor following harvest. Neither treatment showed any negative impact on the overall quality characteristics of ripe fruit, suggesting that they may be practical tools for increasing the ripening rate of Bisir ‘Helali’ dates.     

Factors involved in the production of plantlets from date palm callus cultures

Summary A discussion of a suitable procedure to rapidly propagate free-living date palms (Phoenix dactylifera L.) from callus cultures is presented. Embryogenic callus derived from lateral bud explants was subjected to various auxin treatments in liquid and agar media including p-chlorophenoxyacetic acid. -naphthaleneacetic acid or 2,4-dichlorophenoxyacetic acid at 0.0, 0.1, 1.0 and 10.0 mg/l concentrations in order to obtain optimum growth. Subsequent plantlet initiation from callus was found to be related to initial auxin pretreatments upon subculture to medium devoid of hormones. Plantlet production from callus subcultured from media containing 0.0, 1.0 and 10.0 mg/l auxin concentrations was notably lower than from callus precultured on the 0.1 mg/l auxin levels. In order to improve in vitro adventitious rooting isolated plantlets were cultured on media containing 0.0, 0.1, 1.0 and 10.0 mg/l concentrations of indole-3-acetic acid or -naphthaleneacetic acid in various physical environments. Optimum adventitious rooting responses and survival in free-living conditions were obtained by culturing plantlets in medium containing 0.1 mg/l for 8–16 weeks prior to transplanting to soil. Axillary shoot outgrowths (offshoots) were found to be common in plantlets cultured on a variety of media once an adequate root-shoot system was developed. Mention of a trademark name or proprietary product does not constitute a guarantee or warranty of the product by the US. Department of Agriculture and does not imply its approval to the exclusion of other products that may be suitable.     

Interspecific hybridization between Vigna unguiculata (L.) Walp. and V. vexillata (L.) A. Rich. through in vitro embryo culture

Immature embryos resulting from the cross V. vexillata × V. unguiculata were cultured on MS medium supplemented with 2,4-D (2 mg/l) and resulted in embryogenic calli. Thirteen hybrid regenerants were obtained via organogenesis by subculturing the calli on MS medium supplemented with BAP (2 mg/l) + adenine sulphate (40 mg/l) + CH (500 mg/l) + cowpea tender pod extract (10%). The interspecific regenerants showed intermediate morphological traits between the parents for leaf shape, pod colour and seed coat colour. The hybrid plants inherited stem, leaf and pod hairiness of the wild species which could serve as a mechanical barrier against viral vectors. Electrophoretic studies of two isozyme systems, peroxidase and esterase, also confirms the hybrid nature of the regenerants as they expressed unique bands of both parents. Cytological study of the meiotic chromosomes revealed high frequency of univalent formation in the hybrids suggesting that the genomes of the parental species are structurally differentiated. The hybrid regenerants exhibited high enzyme activity for three enzymes viz., peroxidase, polyphenol oxidase and phenyl alanine ammonia lyase over the cultivated parent which may be useful in conferring resistance against viral pathogens. This revised version was published online in July 2006 with corrections to the Cover Date.     

Morphological variations and isoenzyme polymorphism of date palm clones from in vitro culture acclimatized and established on soil in South Morocco

Five clones of date palm (Phoenix dactylifera L.) corresponding to three reputed Moroccan cultivars (BFG, JHL and BSK) and two selected seedling genotypes (S16 and S35), derived from in vitro culture, were acclimatized and planted in Errachidia soil (South Morocco) in 1989. After 10 years of culture, clones were evaluated on the presence/absence of the two principal morphological characters (offshoots, inflorescences). Then, their leaf extracts were subjected to electrophoresis on polyacrylamide gels in order to determine their isoenzyme polymorphism. Inflorescence formation is relatively more frequent in the clones S16 and S35 than in clones JHL and BSK. Clone BFG showed an intermediate situation. High isoenzyme variations were found for the oxidoreductases (peroxidases (POX) and polyphenoloxidases (PPO)), the transferase (glutamate oxaloacetate transaminase (GOT)) and the hydrolases (esterases (EST) and endopeptidases (ENP)) enzymes. The factorial component analysis exhibited a negative correlation between the two principal morphological characters (presence of inflorescences and offshoots formation). Date palm clones typified respectively by high inflorescences and high offshoots formed two separated groups showing each one some isoenzyme characteristics. All morphological and enzyme variations were discussed in respect to the genotype effect on the micropropagation process.     

颐和园古桑树的组织培养与快速繁殖

以北京颐和园古桑树为外植体进行组织培养,取桑树当年新生嫩茎切段为外植体进行,其中不定芽启动培养基为MS+6-BA 0.5 mg/L+NAA 0.1 mg/L+PVP 0.2 g/L;将无菌苗接到最适分化培养基MS+ 6-BA 0.8 mg/L +NAA 0.1 mg/L +PVP 0.2 g/L上,组培苗分化率高;不定根最适诱导培养基为：1/2MS+NAA 0.5 mg/L +PVP 0.2 g/L,生根率达100%。组培苗移栽成活率达95%。     

In vitro adventitious shoot regeneration for effective maintenance of male sterile niger Guizotia abyssinica (L.f.) Cass.)

A rapid and efficient method of in vitro plant regeneration for large scale propagation of male sterile plants of niger (Guizotia abyssinica) was developed. Leaf segments from mature plants were cultured on Murashige & Skoog's basal medium (MS) supplemented with N⁶-benzyladenine (BA) and kinetin individually and in combination with low concentrations of indole-3-butyric acid (IBA); and -Naphthaleneacetic acid (NAA). Prolific direct adventitious shoot regeneration occurred on most of the media tested. The best response in terms of frequency of shoot regeneration and the number of shoots per leaf explant was observed on medium supplemented with 2.22 M BA. Transfer of shoot bud clusters to fresh medium with same composition promoted further multiplication of the shoot buds, while medium with reduced BA concentration (0.89 M) facilitated shoot elongation. Shoots that were rooted on half-strength MS medium gelled with 0.2 or 0.4% agar and supplemented with 4.9 M IBA survived with a frequency of 61.36% on transfer to ex vitrum conditions. Field evaluation of the regenerants revealed the genetic stability of the plantlets and are being used in breeding of experimental hybrids.     

Genetic Diversity of the Date Palm (Phoenix dactylifera L.) from Algeria Revealed by Enzyme Markers

Analysis of the enzymatic polymorphism of the date palm, Phoenix dactylifera L., was undertaken to evaluate the genetic variability of Algerian cultivars. Seven enzyme systems (alcohol dehydrogenase, diaphorase, aspartate aminotransaminase, acid phosphatase, endopeptidase, leucine aminopeptidase and phosphoglucomutase) were visualized by electrophoresis. Genetic hypotheses appear to show 7 polymorphic loci and 16 alleles were counted. The study of diversity showed a high percentage of polymorphic loci, strong heterozygosity and considerable genetic diversity. The study shows that genetic variability is greater in the cultivars in the western regions (Saoura-Touat) than in the eastern regions (Rhir-Zibans). An identification key was devised and nearly 65% of the cultivars studied were identified from 5 enzyme systems.     

In vitro culture of tissues,cells and protoplasts of Trifolium alexandrinum L. (Egyptian clover)

Summary Callus induction derived from root, hypocotyl and cotyledon explants were investigated for four cultivars of Trifolium alexandrinum L. The Murashige & Skoog (1962) medium containing 2.0 mg/L NAA and 0.5 mg/L BAP was able to induce callus from different explants. A wide range of culture media were tested to determine the morphogenetic potential of different callus types derived from different explant types. Shoot morphogenetic development was observed with the cultivars Sakha 4 and Giza 10. Cell suspension cultures were established from hypocotyl derived callus. Methods for isolation and culture of protoplasts from cotyledons are described.     

Genetic analysis of heading date and other agronomic characters in barley (Hordeum vulgare L.)

Genetic analyses of heading date, tiller number, plant height, grain yield, kernel weight, and plump and thin kernels were made in three six-rowed barley crosses (Hordeum vulgare L.) involving four cultivars. Six populations, P₁ , P₂ , F₁ , F₂ , BC₁ , and BC₂ , from each cross were grown and evaluated at Fargo and Prosper, North Dakota, 1982. Parental means within crosses generally were different except for tiller number. Comparison of generation means suggested that late heading was dominant to early, high kernel weight was dominant to low, and kernel plumpness was influenced by additive gene action. The relationship between yield and heading date was not consistent among crosses and positive r values were quite low. It should be possible to select early maturing, high yielding segregates with plump kernels. Heterosis over the mid-parent was quite similar among crosses for heading date, but there was no heterosis over the high parent. Inbreeding depression was fairly constant for heading date, but was less consistent for yield. The lack of uniformity for estimates of inbreeding depression can be related to environmental variation and to its influence on type of gene action. The ratio of additive to dominance variance was inconsistent among crosses for heading date and yield. These data suggest selection for these characters should be delayed past the F 2 generation. Broad sense heritabilities for heading date ranged from 42 to 86%. Values obtained for grain yield were more consistent among broad sense than narrow sense estimates. Genetic advance estimates were low due to lack of additive variance. This revised version was published online in July 2006 with corrections to the Cover Date.     

Phytotoxic effect of culture filtrate from Fusarium subglutinans the causal agent of fusariose of pineapple (Ananas comosus (L.) Merr

Two pineapple varieties differing in resistance to fusariose were examined for the phytotoxic effect of Fusarium subglutinans culture filtrate. The cultivars were Perolera (more resistant to pathovars of Fusarium subglutinans) and Smooth Cayenne (more susceptible). The phytotoxic effect of culture filtrate was assessed in tissue culture pineapple plantlets (by electrolyte leakage and placing the culture filtrate on wound leaf segments) and callus (inhibition of growth). Smooth Cayenne proved to be the most sensitive cultivar in each test, whereas Perolera showed resistance to the culture filtrate and its callus grew in the presence of high concentrations of culture filtrate that were completely toxic to Smooth Cayenne. These results show that plants can display cellular resistance to the culture filtrate. This revised version was published online in August 2006 with corrections to the Cover Date.     

Current status and future strategies of in vitro culture techniques for genetic improvement of mungbean (Vigna radiata (L.) Wilczek)

Summary Mungbean is an important source of vegetable protein for the growing population in many developing countries of South East Asia. Its production is limited due to its susceptibility to diseases and insect pests besides many other undesirable agronomic traits. Strategies for increasing and stabilising its production have been to develop varieties resistant to diseases, pests and with other desirable agronomic traits. Genetic improvement of this crop by classical breeding has met with limited success due to the lack of sufficient and satisfactory level of genetic variability within germplasm. Recent advances in biotechnology have offered the opportunity to develop new germplasms. The development of such technologies largely depends on efficient regeneration of sexually mature plants from organs, tissues and protoplasts. An overview of plant regeneration by direct or indirect organogenesis and embryogenesis is presented. The use of in vitro and molecular techniques such as somaclonal variation, screening for various desirable traits, interspecific crossing and genetic transformation to supplement conventional breeding, for genetic improvement of this crop is described. The advantages and limitations of these techniques along with directions for future research are discussed.     

Genetic variability in plants regenerated from in vitro culture of sunflower (Helianthus annum L.)

In vitro regenerated sunflower (Helianthus annuus L.) plants (R₁) were self-fertilized and the R₂ generation was evaluated for qualitative traits. A broad range of phenotypic variation was observed and mutation frequencies were calculated. Some in vitro induced variant phenotypes were similar to known spontaneous or induced mutations in sunflower, while others were new. Chlorophyll and carotenoid deficiencies, chimaerical variegation, fasciated stem and capitulum, abnormal shoot development, and other morphological variations, were noted. Substitution of anthers with petaloid structures in a disk-floret mutant indicates a possible homeotic mutation induced by in vitro tissue culture.     

小麦幼胚的脱分化状态及再生性能研究

通过对30个品种（系）的研究，确立了小麦幼胚最佳取材时期的种子形 态指标：在幼种子发育到嫩绿色，脊部由光亮转变为无光泽并具一层绒毛时，幼胚刚好发育到透明的后期和半透明前期。该时期仅有几个小时。探讨了不同基因型的幼胚的致密愈伤组织诱导状况及其继代保持和分化能力，并获得了良好的再生体系。使得致密愈伤组织诱导率达到91％，在14d时分化频率达到96．7％，70d时分化频率仍可达62％。并就温室材料和大田材料以及不同年份大田材料的致密愈伤组织的继代保持和分化能力进行了比较。     

The Growth in Culture of Detached Ovaries of Wheat (Triticum aestivum L. em. Thell)

Viable embryos have been recovered from fertilized wheat (Triticum aestivum) ovaries detached on the day of, or up to 7 days post-, anthesis and cultured aseptically fur up to 18 days. The most significant factors in determining the yield of embryos was the plating density, age and complexity of the explants. 26% of ovaries excised on the day of anthesis produced viable embryos if cultured as pairs of florets. The potential use of detached ovary cultures in gametophyte microinjection experiments, rescuing embryos from wide crosses and in chemically manipulating the early stages of embryo development are discussed.     

Development of elite families in oil palm (Elaeis guineensis Jacq.)

Summary Data from 29 dura female parents, 14 pisifera male parents and their offspring were assembled to obtain quantitative genetic information on optimizing selection of elite families. Phenotypic values of the offspring were partitioned into general combining ability (GCA) of the parents and their specific combining ability (SCA). Parental GCA-values for leaf-Mg content (LMG), height increment (HT), leaf area ratio (LAR), leaf area (LA), frond production (FPR), vegetative dry matter production (VDM), the rate of increase in leaf area against age (k) and maximum leaf area (L_m) were entered in a multiple regression analysis as predictors of oil-and-kernel yield and Harvest Index (HI), i.e. the proportion of dry matter used for oil and kernel production, of the offspring. The analysis revealed that both selection objectives are positively influenced by indirect selection for high LAR and LMG, and for reduced VDM with FPR as main component. But palms with high values for LA, L_m and HT should not be considered to avoid selecting palms partly for their ability to compete with neighbouring palms for light. The contribution of SCA was relatively small for most of the characters studied, except HT, FPR and HI. From the standard deviations for GCA and SCA of the dura and pisifera parents, i.e. _GCA and _SCA, it was extrapolated that selection can potentially enhance bunch yield (by 24%), oil-to-bunch ratio (19%), kernel-to-bunch ratio (29%), oil-and-kernel yield (31%) and HI (56%) above the mean phenotypic value obtained for the offspring of the present investigation. Heritabilities, calculated as the square of the correlation of the GCA-and phenotypic values of the female parents, were low for bunch yield, oil-and-kernel yield and HI, but moderate for LAR, LMG and VDM; levels were high for oil and kernel extraction rate. These outcomes were confirmed by the correlations between values of the female parent and their offspring. Among the unbalanced incomplete mating designs studied, the rectangular lattice and the alpha design are most suitable to compare GCA-values of pairs of parents. A breeding strategy adapting the recently introduced practical possibility of clonal reproduction is discussed.     

西藏青稞冬春性鉴定及抽穗期多样性与稳定性分析

为提高青稞种质资源的利用效率并筛选优异杂交亲本,分别在西藏林芝和拉萨两地的春播和秋播环境下对1605份青稞种质资源进行冬春性鉴定和抽穗期多样性分析。设拉萨春播2个播期,即正常春播I和晚10 d春播II,以2个播期的抽穗期变化作为冬春性区别依据。结果表明,96.2%的西藏青稞地方品种为春性,在西藏3个生态区均有分布,冬性品种仅有3.8%,且主要分布在以林芝为主的藏东南生态区;抽穗期的Shannon-Wiener's多样性分析表明,春播条件下抽穗期多样性高于秋播,拉萨春播条件下抽穗期多样性最高,林芝春播次之,林芝秋播最低;在相同环境下,地方品种的多样性高于育成品种;在环境稳定性分析中, 368份春性地方品种和21份育成品种在不同环境中抽穗期较为稳定,其中康青3号在两地所有参试品种中抽穗期稳定性最高。本研究为全面理解西藏青稞资源的冬春性、抽穗期多样性和环境稳定性提供了参考,为广适应性青稞品种培育筛选出环境稳定性佳的亲本材料。     

In vitro breeding of high-linolenic, doubled-haploid lines of linseed (Linum usitatissimum L.) via androgenesis

In the course of a breeding programme for high-linolenic-acid linseed (oilflax, Linum usitatissimum L.) doubled-haploid lines (DH-lines) of three F₁ hybrids were generated using an anther-culture technique. A total of 82 DH-lines were generated and multiplied in 1993, of which 39 were suitable for testing in a repeated field trial in 1994. Yield, fat content and fatty-acid composition were determined. The yield of the DH-lines was not significantly different from that of respective mid-parent values. Fatty-acid composition and fat content showed remarkable variation, and DH-lines exceeded cross parents in some cases for both fat content and linolenic-acid proportion. Superior DH-lines surpassed mid-parent linolenic-acid-content values by 4.5% and fat content by 1.9%. The results indicate that anther culture is a useful tool in linseed breeding.     

In vitro chromosome doubling with colchicine during microspore culture in wheat (Triticum aestivum L.)

Isolated microspores of two DH lines of wheat were treated with 8 different colchicine concentrations up to 3 mM for either 24 h or 48 h during microspore culture. Untreated control cultures produced on average 220 embryos per spike (100,000 microspores), 68% of the regenerated plantlets were green, and 15% of the flowering plants were fertile. The colchicine treatments had a significant effect on chromosome doubling as measured by the percentage of fertile regenerants. Using colchicine concentrations around 1 mM the percentage of fertile plants among the regenerants was increased up to 53%. The highest number of embryos and regeneration rates were observed after 24 h colchicine treatment, while the highest frequencies of green plants and fertile plants were obtained with 48 h colchicine treatments. The highest number of DH plants per spike was found after treatment with colchicine concentrations of 300 to 1000 μM. Such treatments resulted in an estimated average between the two genotypes of 23 doubled haploid plants per spike. This revised version was published online in July 2006 with corrections to the Cover Date.