The role of mutations in intraspecific differentiation of groundnut (Arachis hypogaea L.)

Summary Based on morphological diversity, cultivated groundnut (Arachis hypogaea L.) is classified into two subspecies (fastigiata and hypogaea) and further into four botanical types (Spanish bunch, Valencia, Virginia bunch and Virginia runner). In a cross between two Spanish cultivars belonging to ssp. fastigiata, a true breeding variant (Dharwad early runner) sharing some characters of both the subspecies was isolated. The variant, on mutagenesis with ethyl methane sulphonate (EMS) yielded a very high frequency of mutants resembling all four botanical types. Some of the mutants produced germinal reversions to Dharwad early runner in later generations indicating genetic instability. While most of the revertants bred true, some of the mutants continued to segregate, wherein each botanical group of mutants produced all other botanical types. A detailed analysis of the breeding behaviour of mutants revealed several unusual features (such as homozygous mutations, mutation outbursts, segregation distortions, somatic mutations and multiple character mutations) that could not be explained through conventional mutation theory. In the light of these findings, the role of mutations in evolutionary differentiation of the crop and the probable mode of their origin have been discussed.     

Simple sequence repeat markers for botanical varieties of cultivated peanut (Arachis hypogaea L.)

Cultivated peanut (Arachis hypogaea L.) consists of six botanical varieties. Identification of DNA markers associated with botanical varieties would be useful in plant genotyping, germplasm management, and evolutionary studies. We have developed 130 simple sequence repeat (SSR) markers in peanut, 38 of which were used in this study because of their ability in detecting genetic polymorphism among 24 peanut accessions. Eight SSR markers were found useful to classify botanical varieties. Among them, six SSR markers were specific to botanical varieties fastigiata and vulgaris, one to botanical varieties hypogaea and hirsuta, and one to botanical varieties peruviana, and aequatoriana. Also, three of them derived from peanut expressed sequence tags (ESTs) were associated with putative genes. As botanical varieties have different morphological traits and belong to different subspecies in A. hypogaea, these markers might be associated with genes involved in the expression of morphological traits. The results also suggested that SSRs (also called microsatellites) might play a role in shaping evolution of cultivated peanut. Multiplex PCR of botanical variety-specific markers could be applied to facilitate efficient genotyping of the peanut lines.     

Genetic variation among Southern African cultivated peanut (Arachis hypogaea L.) genotypes as revealed by AFLP analysis

The amplified fragment length polymorphism (AFLP) technique, employing two different rare cutters, EcoRI and MluI in combination with the frequent cutter MseI, was used to assess genetic diversity and relationships among 21 closely related cultivated Southern African peanut genotypes. A dendrogram was constructed using Jaccard's coefficient and the UPGMA clustering method. Low levels of polymorphism (on average 2.78%) were detected. Results indicated that both EcoRI/MseI and MluI/MseIAFLP enzyme combinations efficiently detected polymorphism within closely related cultivated peanut, although the EcoRI/MseI enzyme combination detected more fragments per primer combination (on average 67.8) as opposed to29.7 by the MluI/MseI enzyme combination. All 21 genotypes could be uniquely distinguished from each other with a minimum of three MluI/MseI primer combinations. Genetic data correlated well with known species and pedigree data, dividing the 21 genotypes into two main groups corresponding to the two subspecies of Arachis hypogaea namely fastigiata and hypogaea. Divisions within the two main groups correlated with botanical types and pedigree data. This is the first report where MluI/MseI primer combinations were used on cultivated peanut and also the first successful detection of polymorphisms between closely related cultivated peanut genotypes worldwide. This revised version was published online in July 2006 with corrections to the Cover Date.     

The classification of varieties of groundnut (Arachis hypogaea L.)

Summary An expanded scheme of groundnut variety classification is presented which is revised in the light of recently collected West African material and is designed in concordance with the most recent taxonomic treatment of the species Arachis hypogaea L. This species consists of two subspecies ssp. hypogaea and ssp. fastigiata Waldron. Each subspecies contains two botanical varieties. Those of ssp. hypogaea are var. hypogaea and var. hirsuta Kohler and those of ssp. fastigiata Waldron are var. fastigiata and var. vulgaris Harz. Each infraspecific taxon contains a morphologically distinct group of cultivated varieties.Grain Legume Research Laboratory, Agricultural Research Council of Malawi, POB215, Lilongwe, Malawi.Department of Agricultural Botany, University of Reading, Reading, England.     

Effects of Growth Regulators and Genotypes on Callus and Embryoid Induction from Maize Anther Culture *

Fifty genotypes and ten growth-regulator combinations were used in two experiments (I and II) to investigate genotype and hormonal effects on production of callus and embryoids via anther culture in maize (Zea mays L.). Hormonal effects across all genotypes were not significant in either experiment. However, highest callus-induction frequencies in both experiments occurred on YP basal medium plus 2,4-D at 2.0 mg 1^?1, and kinetin at 1.5 mg 1^?1 indicating that this combination was more effective than others. Genotypic differences in callus or embryoid induction across all media were significant. The most responsive genotypes in experiment I were single-cross hybrids Yuanwu × 592 and K727 × K305, which produced 18.3 and 6.7 % calli, respectively, with their appropriate media. The most responsive entry in experiment II was CIMMYT Pool 29, which produced 15.0 % calli on appropriate medium and an average of 10.0 % calli across 10 media. Twenty-three plantlets was regenerated from this study. Most of them developed embryogenically.     

Ribosomal DNA repeat unit polymorphism and heritability in peanut (Arachis hypogaea L.) accessions and related wild species

Repeat unit length and restriction site variation in ribosomal RNA geneclusters (rDNA) was surveyed in 77 Arachis accessions, includingsamples from 39 accessions of cultivated Arachis hypogaea(2n=4x=40), 36 accessions representing 15 related tetraploid and diploidwild species, and two synthetic amphidiploids. Total genomic DNA wasdigested with five restriction enzymes, and probed with three heterologousribosomal clones of wheat and broad bean. Four rDNA repeat unit lengthclasses were recognized in the Arachis species. Restriction site analysisshowed that some SacI, BamHI and TaqI cleavage sites in rDNA unit werehighly conserved. With few exceptions, the variable BamHI and EcoRV siteswere able to differentiate the taxonomic sections and species, respectively.Arachis hypogaea and A. duranensis accessions produced fourrDNA length classes. Among these, three were identical with those of otherArachis species. A SacI restriction site (s) from probe (Ver6-5) cangenerally distinguish the two subspecies A. hypogaea ssp. hypogaea and A. hypogaea ssp. fastigiata. Forty nine per centof bands were polymorphic across the A. hypogaea accessionsanalysed. This study does not support A. batizocoi to be a progenitorof A. hypogaea. For the gene array, the contribution from eachparental genome can be detected in the two synthetic amphidiploids.     

Plant regeneration via organogenesis and somatic embryogenesis in callus cultures derived from mature zygotic embryos of leek (Allium ampeloprasum L.)

Summary A high frequency plant regeneration system via organogenesis and somatic embryogenesis was established with callus cultures derived from mature zygotic embryos of different leek genotypes (Allium ampeloprasum L.). Four different callus types with varying morphogenetic potential were obtained. Relatively high concentrations of the auxin 2,4-dichlorophenoxy-acetic acid reduced callus weight and subsequent shoot regeneration and primordia formation of the callus. Shoot regeneration and primordia formation of the callus decreased after prolonged subculture on media containing 2,4-dichlorophenoxy acetic acid. A callus growth period of six weeks on Murashige and Skoog medium with 0.25–0.5 mg l^-1 2,4-dichlorophenoxy acetic acid showed the highest rate of shoot regeneration after transfer of callus to regeneration medium with 1 mg l^-1 kinetin.Differences between leek genotypes in callus type, callus weight, shoot regeneration and primordia formation were observed. Histological observations showed that plant regeneration took place, both via the pathway of somatic embryogenesis and organogenesis.Abbreviation 2,4-D 2,4-dichlorophenoxy acetic acid - MS Murashige and Skoog (1962) medium     

Effect of inbreeding and growth regulators on the in vitro androgenesis of wheatgrass, Agropyron glaucum

The capacity of haploid production by anther culture has been analysed in 42 crossbred genotypes (wild type) and in 32 genotypes from inbreeding generations (I₂-I₃) of wheatgrass, Agropyron glaucum. On potato-2 medium, about 50% of the genotypes investigated were capable of embryoid and callus induction. For the regeneration of haploid plants, five media containing various growth regulators were used. The crossbred genotypes were superior to the inbred lines in all the androgenesis parameters examined.     

Plant regeneration through callus initiation from mature embryo of Triticum

The behaviour of diverse Triticum genotypes in the tissue culture response of mature embryo callus was compared, and factors affecting tissue culture response were studied in this paper. Significant differences were detected in callus induction, embryogenic callus differentiation, plantlet regeneration and culture efficiency when mature embryos of 31 plants of different Triticum species were compared. These were the main wheat cultivars of the Chinese northern winter‐type wheat region and breeding lines (Triticum aestivum L.), durum wheat (Triticum durum Desf.), cultivable emmer wheat (Triticumdicoccum Schuble) and the common wheat progenitors Triticum dicoccoides and Triticum aegilopides. The genotype dependency was particularly high in tissue culture of mature embryos of these Triticum genotypes. The efficiency of induction, differentiation and regeneration of mature embryos callus was high in genotypes selected out. Mature embryo‐derived callus of HB341, TS021, SN2618, T. dicoccum, HB188, and T9817 showed better tissue culture response than the other genotypes. Plantlets can be regenerated from mature embryo‐derived callus of 31 genotypes, saving on growth facility resources and time required for the collection of other explants, and providing a solid basis for the genetic transformation and molecular plant breeding of Triticum plants.     

Phenotypic diversity in cold-tolerant peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.) germplasm

Tolerance to low temperature is an important prerequisite for optimal performance of peanut (Arachis hypogaea L.) in a number of temperate peanut-growing environments. One hundred fifty-eight peanut accessions belonging to five botanical types, known to be tolerant to low temperature (12°C) at germination, were evaluated for phenotypic diversity for 15 morphological traits in the 2001 rainy season and for 15 agronomic and two seed quality traits in the 2001 rainy and 2001/2002 post-rainy seasons. Analysis of data, using the residual maximum-likelihood approach indicated that variance components due to genotypes were significant for all traits in the rainy and for all but two traits in the post-rainy season. Clustering based on scores of nine principle components delineated four clusters. The cold-tolerant genotypes and the standard control cultivars in the four clusters differed in mean, variance, and range both during rainy and post-rainy seasons for a range of agronomic traits, indicating the diversity among the clusters. The cold-tolerant accessions were superior to control cultivars for several agronomic traits compared with their respective controls in both the rainy and post-rainy seasons, so their use in breeding should result in genetically diverse cold-tolerant high-yielding peanut cultivars.     

Somatic in vitro culture response of Lolium perenne L.: genetic effects and correlations with anther culture

Summary Genotypic effects on callus induction and plant regeneration in callus, suspension and protoplast culture, and their correlations with both phenotypic and GCA-values for anther culture response, were studied using 21 genotypes of perennial ryegrass. Differences between genotypes accounted for approximately 40% of the total variation for callus induction and initial callus growth, and 59 and 83% of the variation in callus culture for regeneration percentage and percentage of green plants. Effects of genotypes were less pronounced in suspension culture, where suspensions from the same genotype often behaved differently. Some suspension cultures retained their capacity for green plant regeneration for almost two years, repeatedly producing 80–100% green regenerants during this period. Genotypes with high regeneration percentage and a large proportion of green plants from callus culture were also superior in suspension culture for both regeneration performance and longevity. Regeneration percentage and percentage of green plants were uncorrelated, and probably under different genetic control. While capacity for green plant formation from the different genotypes showed no correlation between anther culture and somatic in vitro culture, a positive correlation was observed between the regeneration percentages in somatic in vitro culture and anther culture (r=0.44^*–0.85^***), suggesting some common genetic control of the two systems.     

Combining ability for yield,protein and oil of peanut lines from South American centers of diversity

Summary Ten peanut (Arachis hypogaea L.) lines, two from each of five centers of diversity in South America, and the F₂ generation of all possible crosses among them were used to characterize the variation for yield, fruit and seed traits and protein and oil content and to determine the breeding potential of the parents. Two of the parental lines belong to subspecies hypogaea and the other eight to subspecies fastigiata.Several crosses gave heterotic responses for yield. The parents of the crosses with significant heterosis generally came from different centers of diversity.Variation among both general and specific combining abilities was significant for all traits with the exception of the SCA estimate for protein percent. The component of variation for GCA was larger than the SCA component for all traits.A Spanish line and a Valencia-like intermediate line from the Guarani region had the highest GCA effects for fruit yield, seed yield and meat content and transmitted consistently their characteristics to their F₂ progeny. Some lines showed significant but small maternal effects for oil, protein and oil + protein percent. Differences between reciprocal crosses were significant for fruit length, fruit weight and oil + protein percent.A significant portion of the variation among crosses was explained by differences among parental lines within centers of diversity. This supports the fact that centers are founded because of the diversity of the peanuts grown there.The performance of parental lines per se was useful in predicting the performance of the lines in hybrid combination for all traits except fruit yield.Paper number 6141 of the Journal Series of the North Carolina Agricultural Research Service (USA). This work was partially supported by CSRS Research Agreement 701-15-51.     

Plant Regeneration from Protoplasts in Linum*

Protoplasts of eight wild Linum species and 14 genotypes of flax, isolated from different parts of seedlings and from shoot apices, were regenerated to callus in medium V-KM (Binding and Nehls 1977). Plant regeneration was obtained in L. alpinum, L. amurense, L. hologynum, L. perenne, L. salsoloides and eight genotypes of L. usitatissimum on agar media B5 (Gamborg et al. 1968), B5C (Binding et al, 1981), MS (Murashige and Skoog 1962) and MSC (MS with 5% coconut endosperm) containing 2.5μM 6-BA. An adventitious embryo was regenerated in L. alpinum. Callus differentiation like sponge parenchyma, was observed in protoplast-derived tissues of L. usitatissimum. Shoots organized roots under slightly divergent conditions in the different species.     

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.     

Phenylacetic acid stimulation of direct shoot formation in anther and somatic tissue cultures of rice (Oryza saliva L.)

The effect of phenylacetic acid (PAA) on rice (Oryza saliva L.) anther culture was investigated with six genotypes, using 2,4-D as control. In the two-step culture protocol, replacing 2, 4-D with PAA in the induction medium did not influence callus induction but significantly improved the shoot differentiation from callus, particularly in the indica cultivar Teqing. The anther-derived calli of all genotypes regenerated shoots directly on the callus induction medium containing PAA. Most of the directly-regenerated plantlets had well-developed root systems and were therefore readily transplanted into soil. The improved shoot differentiation potential and the frequency of direct regeneration depended on genotype, basal medium and PAA concentration. The one-step green shoot regeneration frequencies obtained were 1.98% with the indica cultivar ‘129’, 1.5% with the indica × japonica hybrid ‘Teqing/02428’ (F₁), and 1.98% with the indica × indica hybrid ‘Waiyin 2/C.B.’ (F₁). The PAA-based one-step method was most effective on the anther culture of indica genotypes. Three DH populations have been constructed from hybrids (F₁) via one-step culture. PAA also enhanced the one-step plantlet formation in rice somatic tissue culture.     

Profiling peanut (<Emphasis Type="Italic">Arachis hypogea</Emphasis> L.) accessions and cultivars for oleic acid and yield in Botswana

The ratio of monounsaturated oleic fatty acid to polyunsaturated linoleic fatty acid (O/L) and the associated agronomic traits were profiled for local peanut (Arachis hypogaea L.) cultivars and accessions in Botswana. The research was tested on 16 entries planted in four localities across the country. The average total oil yield of the genotypes studied was about 42.1% and there was no significant difference in total oil yield (P < 0.05) across the four research sites and among all entries (accessions). The average mean O/L ratio for all entries was about 1.47. The highest was that of the locally improved variety, Peolwane (2.58), which was significantly different (P < 0.05) from all other accessions, and the lowest is that of the local accession GO 63 (1.23). This study also profiled the agronomic yield traits to identify genotypes that have both high oil quality and high yielding potential. No correlation was detected between pod yield and O/L ratio among the tested genotypes. The station with little rainfall but adequate repartition showed a very high O/L ratio for some genotypes, and future work is recommended to test this interesting observation on the correlation between moisture stress and oil quality.     

Plant regeneration from embryo-callus culture in barley

Summary Plants were regenerated from callus cultures initiated from immature embryos of barley, Hordeum vulgare L. Immature embryos from seven diverse genotypes were cultured on modified Murashige and Skoog (MS) medium supplemented with 1.5 mg 2,4-D and 6.5 mg IAA/l. Of the 249 embryos cultured, 30% initiated callus within 8 days. Subculture of callus for 80 to 100 days on half-MS medium supplemented with 0.5 mg/l 2,4-D and 1.0 mg/l zeatin resulted in organogenesis. Culture of organogenic calli for 30 days on half-MS medium without growth regulators produced plants which originated mostly via multiple shoot formation. Callusing response of the tested genotypes ranged from zero to 44%; however, only 23% of the calli were regenerative. Regenerated plants included variants for chlorophyll deficiency, plant height, stem thickness, spike shape, pollen fertility, seed set and ploidy.     

Genetic Variation for Anther Callus Inducibility in Crosses of Highly Culturable Winter Wheats

Heritability studies suggest that variation among genotypes for callus inducibility should respond to selection. The objective of this study was to determine if crosses among parents with high callus initiation frequencies would produce segregation from which genotypes with improved callus inducibility could be selected. The winter wheat (Triticum aestivum L.) cultivars‘Centurk 78’, ‘Norstar’ and ‘Norwin’ were chosen as parents, based on their high callus initiation frequencies and diverse parentage. F₂ populations from crosses between each of these parents were evaluated for callus initiation frequencies and callus vigor as determined by growth rate and healthy appearance. Heterosis was detected for callus initiation in the‘Norwin’בNorstar’ cross and for callus vigor in all crosses. Broad sense heritability estimates of 0.25 and 0.37 for callus initiation in the ‘Centurk 78’בNorstar’ and ‘Norwin’בNorstar’ crosses, respectively, suggest this trait can be improved by selection. Heritability estimates for callus vigor were less promising, ranging from 0.11 to 0.24.     

抗青枯病花生种质的遗传多样性

以栽培种花生2个亚种4个植物学类型的31份对青枯病具有不同抗性的种质为材料,通过SSR和AFLP技术分析了其DNA多样性,并与通过形态和种子品质性状揭示的表型多样性进行了比较。结果表明,不同类型的抗青枯病花生品种之间存在丰富的DNA多样性,SSR揭示的品种间遗传距离大于AFLP揭示的品种间遗传距离,基于二者的聚类分析结果趋势一致,结合花生的植物学类型、地理来源和系谱分析,以SSR的聚类结果与表型性状的聚类结果更为吻合。感病优质高产品种“中花5号”与密枝亚种的普通型和龙生型的抗病材料的差异很大,与育种中被广泛利用的抗源“协抗青”和“台山三粒肉”的差异相对较小,与“远杂9102”的差异更小。     

2种甜玉米自交系幼胚再生体系的建立

本试验研究不同基因型、幼胚大小、培养基类型及培养基添加物对2种甜玉米(2-105和2-118)自交系幼胚愈伤及胚性愈伤组织诱导率的影响。结果表明,基因型、培养基类型、幼胚大小及不同浓度脯氨酸和2,4-D处理对愈伤及胚性愈伤诱导率影响差异均显著。2种基因型最适宜的基本培养基为N6培养基,且2-118的愈伤及胚性愈伤诱导率显著高于2-105。幼胚在1.5~2.0 mm时,2-105和2-118的愈伤及胚性愈伤诱导率均最高,分别在77%和51%以上。2-105和2-118在脯氨酸浓度分别为650 mg/L、750 mg/L时幼胚的愈伤及胚性愈伤诱导率最高,且显著高于其它2个水平。2种基因型在2.5 mg/L的2,4-D诱导下愈伤及胚性愈伤诱导率均最高,且均显著高于其它2个水平。本研究获得了2种甜玉米自交系的再生体系,为甜玉米遗传转化和分子育种提供基础。