In vitro organogenesis and plant formation in Vigna radiata (L.) Wilczek

In vitro organogenesis was achieved from calluses derived from cotyledon and hypocotyl explants of Vigna radiata on MS medium. Organogenic calluses were induced from both cotyledon and hypocotyl explants excised from 3-day-old seedlings on MS medium containing NAA (1.07 m and BA (2.22 m) and 2,4-D (0.90 m) and BA (2.22 m) combinations respectively. Regeneration of adventitious shoots from cotyledon derived callus was achieved when they were cultured on MS medium supplemented with NAA (1.07 m), BA (8.88 m) and 10% coconut water. Hypocotyl derived calluses produced adventitious shoots when cultured on MS medium fortified with BA (6.66 m), TDZ (2.5 m) and 10% coconut water. Addition of GA at 1.73 m favored maximum 3 elongation of shoots. Regenerated shoots produced prominent roots when transferred to half strength MS medium supplemented with 4.90 m IBA. Rooted plantlets, thus developed were hardened and successfully established in field. Among the different carbohydrates and media tested, 87.64 m sucrose and MS+B5 medium proved best for maximum production of shoots. This protocol produced an average of seven plants per hypocotyl derived callus and 15 plants per cotyledon derived callus over a period of 3 months.     

Alleviation of fungicide-induced phytotoxicity in greengram [Vigna radiata (L.) Wilczek] using fungicide-tolerant and plant growth promoting Pseudomonas strain

This study was designed to explore beneficial plant-associated rhizobacteria exhibiting substantial tolerance against fungicide tebuconazole vis-à-vis synthesizing plant growth regulators under fungicide stressed soils and to evaluate further these multifaceted rhizobacteria for protection and growth promotion of greengram [Vigna radiata (L.) Wilczek] plants against phytotoxicity of tebuconazole. Tebuconazole-tolerant and plant growth promoting bacterial strain PS1 was isolated from mustard (Brassica compestris) rhizosphere and identified as Pseudomonas aeruginosa following 16S rRNA gene sequencing. The P. aeruginosa strain PS1 solubilized phosphate significantly and produced indole acetic acid, siderophores, exo-polysaccharides, hydrogen cyanide and ammonia even under tebuconazole stress. Generally, tebuconazole at the recommended, two and three times the recommended field rate adversely affected the growth, symbiosis, grain yield and nutrient uptake in greengram in a concentration dependent manner. In contrast, the P. aeruginosa strain PS1 along with tebuconazole significantly, increased the growth parameters of the greengram plants. The inoculant strain PS1 increased appreciably root nitrogen, shoot nitrogen, root phosphorus, shoot phosphorus, and seed yield of greengram plants at all tested concentrations of tebuconazole when compared to the uninoculated plants treated with tebuconazole. The results suggested that the P. aeruginosa strain PS1, exhibiting novel plant growth regulating physiological features, can be applied as an eco-friendly and plant growth catalyzing bio-inoculant to ameliorate the performance of greengram in fungicide stressed soils.     

Chlorophyll mutations in mungbean (Vigna radiata (L.) Wilczek)

Summary Twenty mutants isolated from Latisail, Jhingasail and Pankaj varieties of rice (Oryza sativa L.) were screened for two aspects of nutritive quality, namely crude protein content and distribution pattern of protein in the endosperm. Observations revealed a wide variation for both characters, and while there was no consistent association between protein content and test grain weight, which varied between varieties, a positive correlation between protein content and grain sterility was noted. In a few mutants protein distribution was observed to be varied and showed a similarity to optimum milling characteristics.     

Comparative efficacy of pedigree selection and selective intermating in greengram (Vigna radiata (L.) Wilczek)

Summary Selective intermating and pedigree selection methods were applied simultaneously to highly heterogeneous and heterozygous base populations of greengram in order to compare their relative efficacy in terms of evolving the number of productive lines as well as their production potential. Selection after two cycles of selective intermating was found to be a better method than traditional pedigree selection. The demerits of pedigree selection and merits of selective intermating are discussed. It is suggested that selective intermating replace the widely adopted but less effective pedigree selection for generating promising new material in such autogamous crops as greengram.Part of Ph.D. Thesis submitted by the senior author to the Haryana Agricultural University, Hisar, India     

Culture conditions effecting plant regeneration from cotyledons of Vigna radiata (L.) Wilczek

Conditions for plant regeneration from excised cotyledons of Vigna radiata were studied. Complete plant developed from the uncallused proximal ends of cotyledons on Murashige & Skoog's (MS), Gamborg's (B₅) and C (MS salts + B₅ vitamins) basal media. The basal medium C was found to be best for plant regeneration. Regeneration frequency, however, varied with genotype, size, orientation and age of explant and the different plant growth regulators combination in the medium. Addition of cytokinins induced callusing at the proximal ends of cotyledons followed by multiple shoot formation. Out of 6-benzyl aminopurine (BAP), kinetin (KIN), N (^–2 isopentyl) adenine (2iP) and adenine sulphate (AS), only BAP and KIN were found to be more effective in enhancing the frequency of shoot regeneration. BAP at 1×10^-1M induced maximum (60%) shoot regeneration whereas maximum number of shoots (8 to 9 shoots) per explant was observed with 5×10^-6M BAP. Cotyledons excised from two-day old seedlings were most regenerative. The regenerative response of cotyledons decreased when sliced into two equal parts either longitudinally or transversely. Callusing and organogenic differentiation occured only if the petiolar end of cotyledons was in contact with medium. None of the tested treatments were effective in inducing shoot bud differentiation from subcultured callus. Well developed shoots rooted when incubated on half strength MS, MS and MS basal medium supplemented with IAA (5×10^-6M). The rooted plants were transferred to pots and later established in the field with 60% success.Abbreviations AS adenine sulphate - BAP 6-benzylaminopurine - B₅ medium after Gamborg et al. [6], - C Medium with MS salts + B₅ vitamins - 2iP N (^–2 isopentyl) adenine - IAA indole-3-acetic acid - KIN Kinetin - MS medium after Murashige & Skoog [21] - NAA 1-napthaleneacetic acid     

Two pathogenesis-related peroxidases in greengram (Vigna radiata (L.) wilczek) leaves and cultured cells induced by Macrophomina phaseolina (Tassi) Goid. and its elicitor

An elicitor has been isolated from Macrophomina phaseolina, the root rot and leaf blight pathogen of greengram. Suspension-cultured cells of greengram were established which responded to the fungal elicitor. When greengram leaves were inoculated with M. phaseolina two new peroxidases appeared. Similarly, two new peroxidases could be detected in suspension-cultured greengram cells when treated with the fungal elicitor. These peroxidases were purified by column chromatography and their molecular masses were 27 and 38 kDa. The new peroxidases detected in both leaves and cultured cells appear to be similar with the same molecular weights.     

Somatic embryogenesis in Vigna radiata (L.) Wilczek.

Somatic embryogenesis was achieved from immature cotyledon derived callus of mungbean, V.radiata (L.) Wilczek in MS liquid medium. Embryogenic callus was induced on MS medium with NAA (5 mg/L). Differentiation of somatic embryos was observed when embryogenic callus was transferred to MS liquid medium containing 2,4-D (1.5 mg/L) and L-proline (50 mg/L). The torpedo shaped embryos were transferred to MS liquid medium with BAP and ABA (1 mg/L each) for maturation and germination. Fifty per cent of torpedo shaped embryos were converted into tiny plants (8-9 plants out of 17) after one week of culture. The germinated embryos were isolated and transferred to MS half strength basal (solid) medium for further development.     

Genetic basis of plant height and its degree of indetermination in mungbean (Vigna radiata (L.) Wilczek)

The genetic basis of plant height at various growth stages and the degree of indetermination of plant height in six mungbean genotypes (NM 92, 6601, NM 89, VC 1560D and VC 3902A) were assessed through half diallel cross. Cultivars, 6601 and NM 92, were the best general combiner for pre-flowering dry matter accumulation and minimum increase in plant height from first flower to 90% pods maturity, respectively. For these traits, the combination NM 92 x NM 89 was the best specific combiner of all the crosses. Both additive and dominant gene effects controlled the inheritance of plant height at first pod and to 90% pods maturity, degree of indetermination of plant height (DDh) from first flower to first pod maturity (DDh1), DDh from first flower to 90% pods maturity (DDh2) and DDh from first pod maturity to 90% pods maturity (DDh3). Plant height at first flower was additively inherited. The additive gene action was predominant as compared to dominant gene action for all the traits examined. High narrow and broad sense heritability estimates for DDh2 showed that better response to selection is possible for the development of mungbean genotypes with minimum increase in plant height during post-flowering development.     

Diversity in greengram (Vigna radiata (L.) Wilczek) germplasm collection and its potential use in crop improvement

Greengram or mungbean (Vigna radiata (L.) Wilczek), which originated in the Indian sub-continent, is an important pulse crop in India. Genetic diversity and patterns of variation among 111 accessions representing the better agronomic types from the entire collection of the National Bureau of Plant Genetic Resources were studied. Wide variation in morphological and agronomic traits viz. number of pods per plant, yield per plant, 100-seed weight, fruit setting capacity, flowering period, maturity, number of pod bearing peduncles, plant height, number of primary branches, length of branch, nodulation and leafiness was observed. Grain yield was found to be significantly correlated with 100-seed weight and pod length. The data were subjected to cluster and principal components analysis. The accessions were grouped into six discrete and well-defined clusters. The study demonstrated the patterns of variation at the population level. The multivariate analysis was useful in identifying a group of accessions with yield enhancing traits within a highly diverse group of accessions and their potential value in greengram improvement is suggested.     

New microsatellite markers isolated from mungbean (Vigna radiata (L.) Wilczek)

In this study, we reported the isolation and analysis of new polymorphic microsatellites in mungbean (Vigna radiata (L.) Wilczek). Twelve out of 210 primer pairs screened in 30 mungbean accessions gave polymorphism. The polymorphic markers detected two to three alleles per locus with an average of 2.08. Observed heterozygosity varied from 0 to 0.133, while expected heterozygosity ranged from 0.095 to 0.498. Tests for Hardy-Weinberg equilibrium (HWE) and pairwise linkage disequilibrium of the polymorphic loci revealed that all loci except MB-SSR14 significantly departed from HWE and four pairwise combinations, viz. MB-SSR14 vs. MB-SSR42, MB-SSR42 vs. MB-SSR87, MB-SSR114 vs. MB-SSR121, and MB-SSR175 vs. MB-SSR231 significantly deviated from linkage disequilibrium. The markers are being used to study genetic diversity and genome mapping of mungbean.     

Differential antioxidative responses of ascorbate-glutathione cycle enzymes and metabolites to chromium stress in green gram (Vigna radiata L. wilczek) leaves

Chromium-induced antioxidative responses of ascorbate-glutathione cycle enzymes and metabolites in green gram(Vigna radiata L. Wilczek) leaves were investigated in both dose and time-dependent manners. Rapid uptake of Cr was observed immediately after the start of treatment. Significant reduction was observed in leaf biomass under 300 μM Cr-treatment. Treatment with 300 μM Cr increases the content of hydrogen peroxide and Superoxide dismytase activity upto initial 96 h, and then gradually declined to the basal level. Ascorbate peroxidase and guaiacol peroxidase activities were low in 300 μM Cr-treated leaves during the first 96 h, but significantly increased therefore, suggesting that increased enzyme activities would be responsible for the removal of H₂O₂. Catalase activities were always suppressed under Cr stress. Contents of reduced ascorbate and dehydroascorbate were significantly decreased under 300 uM Cr-treatment. The reduced glutathione content decreased at early stages of Cr-treatment. However, it was restored to the normal level as in controls thereafter. In contrast, the glutathione disulphide content showed a progressive increase during the initial hours of Cr-treatment. The non-protein thiol content was shown to increase during the first several hours, but it declines at later stages. The present results demonstrate that Cr-induced oxidative stress is an important component of the plant’s reaction to toxic levels of Cr.     

绿豆基因组研究进展

绿豆是亚洲国家重要的经济作物。绿豆基因组的研究工作已开展多年,至今已经发布了6张遗传连锁图谱,然而还未有一张图谱的连锁群数与绿豆(2n=2x=22,n=11)的染色体基数一致。近年来,豆科植物比较基因组学的研究成果,为绿豆遗传连锁图谱的发展提供了新的思路。通过将绿豆遗传连锁图与其他豆类连锁图比较发现,绿豆与小豆、豇豆、普通菜豆、大豆、藊豆以及豆科模式植物—蒺藜苜蓿的基因组间有不同程度的保守性,其中尤以绿豆与普通菜豆基因组间共线性水平高。本文分别从绿豆遗传连锁图谱构建、比较基因组作图以及抗豆象基因定位等方面进行了综述,以期为绿豆遗传研究工作者提供参考。     

Combat of iron-deprivation through a plant growth promoting fluorescent Pseudomonas strain GRP3A in mung bean (Vigna radiata L. Wilzeck)

Microorganisms and plants sustain themselves under iron-deprived conditions by releasing siderophores. Among others, fluorescent pseudomonads are known to exert extensive biocontrol action against soil and root borne phytopathogens through release of antimicrobials and siderophores. In this study, production and regulation of siderophores by fluorescent Pseudomonas strain GRP3A was studied. Among various media tested, standard succinate medium (SSM) promoted maximum siderophore production of 56.59 mg l(-1). There were low levels of siderophore in complex media like King's B medium, trypticase soya medium and nutrient medium (41.27, 29.86 and 27.63 mg l(-1)), respectively. In defferrated SSM, siderophore level was quantified to be 68.74 mg l(-1). Supplementation with iron (FeCl3) resulted in decreased siderophore levels depending on concentration. Siderophore production was promoted by Zn2+ (78.94 mg l(-1)), Cu2+ (68.80 mg l(-1)) whereas Co2+ (57.33 mg l(-1)) and Fe3+ reduced siderophore production (37.44 mg l(-1) as compared to control (55.97 mg l(-1)). Strain GRP3A showed plant growth promotion under iron limited conditions.     

Histological and scanning electron microscopic observations on plant regeneration in mungbean cotyledon (Vigna radiata (L.) Wilczek) cultured in vitro

Explanted cotyledons of mungbean Vigna radiata (L). Wilczek, variety Pag-asa-1, regenerated shoots directly from the basal adaxial side of the petiolar residue on MS medium supplemented with 8.9 M 6-benzyladenine. A simplified and rapid procedure for glycol methacrylate sectioning for histological observations was used to observe shoot initiation. At the time of culture, comparatively smaller and differentially stained epidermal cells were present on the basal adaxial region of the petiolar residue. A meristematic cell mass that developed at 48 h after culture appeared to be of epidermal and subepidermal cell origin. Scanning electron microscopy revealed shoot primordia and approximately 2 nodules at the base of the petiole as early as 48 h after culture. All of these structures developed into shoots during incubation.Abbreviations FAA formalin 5%–70% ethanol, 90%-acetic acid 5% - GMA glycol methacrylate - BA 6-benzyladenine     

Plant regeneration from cotyledonary node explants of mungbean (Vigna radiata (L.) Wilczek)

Sexually-mature mungbean (Vigna radiata (L.) Wilczek) plants were efficiently regenerated from cotyledonary node explants. The explants were capable of directly developing multiple shoots on basal media devoid of any growth regulators. The shoot multiplication was influenced by media composition, growth regulators, age of donor seedling and explant type. The explants with both the cotyledons attached to the embryonic axis excised from 4-d-old seedlings, produced the highest number of shoots (5 or 6) in 100% of the cultures within 2 weeks on B₅ basal medium (BBM) containing BAP or 2-iP, respectively, (at 5x10^–7M) and 3% sucrose. Shoots elongated and developed better using BAP. Increasing micronutrients, carbohydrate and nitrogen levels in the medium above the original formulation of B₅ basal medium appeared to be of no benefit for increasing the number of shoots. The shoots were rooted on basal MS medium or MS containing 10^–6 of NAA, IAA or IBA. This protocol was found applicable to six other cultivars of mungbean. One hundred rooted shoots were successfully established in soil in the glasshouse, where 90% of them survived. The regenerated plants flowered precociously, but produced normal pods and viable seeds.Abbreviations BAP 6-benzylaminopurine - KIN kinetin - 2-iP 6- — -dimethylallyl aminopurine - AdS adenine sulphate - IAA indole-3-acetic acid - IBA indole-3-butyric acid - NAA 1-naphthalene acetic acid - MS Murashige and Skoog (1962) medium - B₅ Gamborg et al. (1968) medium - C medium MS salts + B₅ vitamins     

Interspecific hybridization between Vigna radiata (L.) Wilczek and V. glabrescens

Summary Interspecific hybrids of the mungbean, Vigna radiata (L.) Wilczek (2n=22) and V. glabrescens (2n=44) were generated with the aid of embryo culture. V. glabrescens x V. radiata hybrids were recovered via germination of the immature embryos. Reciprocal hybrids were obtained via shoot formation from embryonic callus. The authenticity of the hybrids was determined by morphological characteristics, chromosome number, and isozyme patterns. The hybrids were highly sterile upon selfing, but backcrossing to the diploid parent yielded viable seeds. Some of the plants resembled the diploid parent morphologically while others resembled neither parent. The backcross plants were sufficiently fertile to give a large number of mature, selfed seeds. Plants obtained differed morphologically and in their isozyme patterns from either parent, indicating introgression. These progeny populations will be used as bridging materials to transfer pest resistance from the wild tetraploid to the cultivated mungbean.     

Thidiazuron-induced high-frequency axillary and adventitious shoot regeneration in Vigna radiata (L.) Wilczek

Summary Prolific shoot regeneration was achieved in mungbean Vigna radiata (L.) Wilczek from 3-d-old in vitro cotyledonary node and hypocotyl explants from seedlings derived from mature seeds on Murashige and Skoog (MS) medium supplemented with thidiazuron (TDZ) (0.9 μM). An initial exposure to TDZ for 20 d and three successive transfers to fresh medium with reduced thidiazuron levels (0.09 μM) resulted in the regeneration of 104 shoots/explant from the cotyledon and 30 shoots/explant from the hypocotyl. Thidiazuron-associated abnormalities such as short compact shoots, fasciation and leaf growth in the form of rosettes were observed in shoots regenerated from hypocotyl explants. Both axillary and adventitious shoot formation from the explants were confirmed by histology. Through repectitive cycles of regeneration in the presence of TDZ, the number of shoots that could be obtained from the two explant classes within 80 d was significantly higher than with previous reports in mungbean     

The wild ancestors of urid and mung beans (Vigna mungo (L.) Hepper and V. radiata (L.) Wilczek)

The crop plants Vigna mungo (urid, urd or black gram) and V. radiata (mung bean or green gram: and related taxa have been studied by seed protein electrophoresis, leaf phenolics chromatography, vegetative morphology, and seed testa patterns. The results disprove the theory that these species are very closely related and have evolved from a single wild taxon. The present evidence shows that V. mungo var. mungo and I. radiata var. radiata have independent lineages and were domesticated from two very distinct taxa. namely V. mungo var. silvestris and V. radoata var. sublobata respectively.     

Chromatin differentiation between Vigna radiata (L.) R. Wilczek and V. unguiculata (L.) Walp. (Fabaceae)

A comparative chromosomal evaluation was carried out between Vigna unguiculata (cowpea) and V. radiata (mung bean) with chromomycin A₃ (CMA₃)/4’,6-diamidino-2-phenylindole (DAPI) banding and fluorescent in situ hybridization (FISH) using 5S/45S ribosomal DNA (rDNA) probes. Both species had symmetric karyotypes (2n = 22), with prevalence of centromeres in chromosomes at median (m) and submedian (sm) regions and chromosomes ranging in size from 2.1 to 1.25 μm (V. unguiculata) and 2.18 to 0.93 μm (V. radiata). Three different banding patterns were identified for V. unguiculata: CMA₃⁺/DAPI⁰, CMA₃⁺⁺/DAPI^–, and CMA₃⁺/DAPI^–. The CMA₃⁺/DAPI⁰ bands were observed in the pericentromeric regions of all chromosomes, while the CMA₃⁺⁺/DAPI^– and CMA₃⁺/DAPI^– bands were co-localized with the 45S rDNA in the subtelomeric position (chromosomes B, G, and D, J, respectively) and in the proximal position in chromosome F. Two pairs of chromosomes (D and I) bearing interstitial 5S rDNA have been also identified. Vigna radiata displayed CMA₃⁰/DAPI⁺ bands distributed in the centromeric region of chromosomes B, C, and F, while CMA₃⁺⁺/DAPI^– bands were co-localized with the 45S rDNA sites in the subtelomeric position of the short arm in the F and K chromosome pairs. Three pairs of 5S rDNA sites were identified, the first in the proximal region of the long arm in chromosome E and the two others in the proximal and subterminal positions in the long arm of chromosome J. These data highlight some divergences regarding the amount and composition of the heterochromatin in both species, allowing the identification of individual chromosomes in V. unguiculata and V. radiata, and a comparison with other members of the Phaseoloid clade.