Somatic Embryogenesis and Plant Regeneration from Leaf Callus of Hordeum vulgare

Explants obtained from the basal portion of leaves of Hordeumvulgare (cv. Karan 92) gave rise to callus when cultured onMurashige and Skoog (MS) basal medium supplemented with 2, 4-dichlorophenoxyaceticacid (2, 4-D). Initially, the callus was friable, shiny-whiteand watery but subsequently some compact, nodular callus appeared.The latter were cultured on MS medium containing 0.05 mg l^–12, 4-D and 0.1 mg l^–1 N6-furfurylaminopurine (kinetin),when plantlets were generated. Histological studies showed thatplantlet regeneration occurred by the formation of somatic embryos.The regenerated plants had the normal diploid chromosome number(2n = 14). Hordeum vulgare, barley, somatic embryogenesis, tissue culture, plant regeneration     

Plant Regeneration Through Somatic Embryogenesis in Leaf Derived Callus of Plumbago Rosea

Regeneration of Plumbago rosea L., a rare medicinal plant, via somatic embryogenesis in callus cultures derived from leaf explants was described. Optimum callus formation was achieved on semi-solid Murashige and Skoog (MS) medium supplemented with 0.25 mg dm^–3 kinetin and 2.0 mg dm^–3 1-naphthaleneacetic acid (NAA). Somatic embryogenesis was achieved upon transferring the 4-week-old callus to a medium containing 1.0 mg dm^–3 kinetic (Kn), 0.5 mg dm^–3 gibberellic acid (GA₃) and 0.1 mg dm^–3 NAA. Embryo maturation and germination was achieved on the half-strength MS basal salts supplemented with 0.01 – 0.25 mg dm^–3 Kn and 2 % (m/v) saccharose. An average of 50 – 60 plantlets were obtained from 150 mg of embryogenic callus within 4 week of subculture. Out of the 50 plantlets about 28 survived in the greenhouse.     

Requirement of Ethylene for Growth of Callus and Somatic Embryogenesis in Medicago sativa L.

The role of ethylene in the growth of callus and somatic embryogenesisin Medicago sativa was examined. The application of 2,5-norbornadiene,a competitive inhibitor of ethylene action, during a 10 d inductionperiod to medium containing 2,4-D and kinetin inhibited thegrowth of callus but did not affect somatic embryogenesis, nordid it affect ethylene production during the induction stage.The exposure of tissue, incubated on differentiation medium,without hormones, to an atmosphere of 2,5-norbornadiene, inhibitedboth growth and embryo maturation and stimulated pigmentation.The inhibition of embryo maturation was observed even in thepresence of norbornadiene at a concentration which did not affectgrowth of tissue. It is suggested that the action of endogenous ethylene is necessaryfor the growth of the callus and embryo maturation. Key words: Medicago sativa, ethylene, callus growth, somatic embryogenesis     

Differentiation in Callus Cultures of Leaf of Two Species of Trigonella

Callusing of leaf segment of Trigonella corniculata L. and Trigonelta foenum graecum L. was observed on Murashige and Skoog's (MS) medium and White's (W) medium supplemented with naphthalene acetic acid (NAA; 0.5 mg/1) and coconut milk (15% v/v). Root, shoot and development of isolated leaf occurred in the calli of both the species on MS medium containing NAA amd coconut milk. Omission of coconut milk with subsequent addition of casein hydrolysale (CH) increased the number of differentiated organs per callus. In both the materials higher differentiation rate was observed at low irradiance than at high irradiance or in complete darkness. Initially the cells were diploid but at the 5th passage the calli became mixoploid wilh predominantly diploid cell populations. The differentiated organs were diploid.     

Thidiazuron对苜蓿愈伤组织的乙烯生成及其体细胞胚胎发生的影响

苜蓿叶柄在Ｂ５Ｈ培养基上诱导并继代所形成的是无色、松软的胚性愈伤组织,内含许多有分生能力的细胞团。当Ｂ５Ｈ中的ＫＴ用ＴＤＺ替换时,愈伤组织变绿变硬,出现管状分子和芽原基的分化,其体胚发生能力丧失,而且其乙烯生成和ＰＡＬ活性升高。乙烯形成酶的抑制剂ＣｏＣｌ２可以抑制ＴＤＺ的上述作用,部分地恢复愈伤组织的体胚发生能力。ＴＤＺ对愈伤组织的这些作用可能与其促进乙烯生成有关。     

Effect of 2-Thiouracil on Cell Differentiation and Leaf Morphogenesis in Cannabis sativa

When supplied through the leaf at low dosage levels, the pyrimidineanalogue, 2-thiouracil, progressively arrests apical growthand organogenesis in young plants of Cannabis sativa. Duringthe treatment period, a generation of leaves arises in whichcell differentiation is inhibited without a parallel reductionin laminal expansion. With the withdrawal of treatment, apicalgrowth is resumed, but severe morphogenetic defects later becomeevident in leaves developing from primordia initiated duringthe treatment period or shortly afterwards, and there is simultaneouslya loss of apical dominance and of the control of phyllotaxis.Growth of a more normal pattern is, however, gradually restoredat stem apices, although local anomalies in the anatomy of thelamina recur throughout the life of the plant, and otherwisenormal leaves continue to show sporadic flecks or streakingsof abnormal pigmentation. The latter phenomenon suggests thatthe analogue induces a persistent state of instability in someof the proplastids of meristematic tissues.     

Differentiation of Nigella sativa seeds from four different origins and their bioactivity correlations based on NMR-metabolomics approach

Authenticity and quality coherence are the major elements in ensuring the consistency of the expected beneficial outcomes from the use of traditional or herbal remedies. Metabolomics offers the possibility of addressing these issues. Principal Component Analysis (PCA) was applied to select the best solvent system for sample extraction. Partial Least Square (PLS) regression analysis was found useful in evaluating the relationship between Nigella sativa seeds from four different origins on the basis of their metabolite profiles. In this study, different bioactivities were displayed by different samples with the Qasemi and Syrian samples exhibited high α-glucosidase inhibitory activity, which was correlated to the high fatty acid contents based on the PLS model. The Ethiopian sample exhibited high DPPH radical scavenging and nitric oxide (NO) inhibition activities, which may be related to the presence of high levels of thymoquinone and thymol. The method was also successfully used to classify “new test” samples into their proper groups.     

Nigella sativa attenuates myocardial ischemic reperfusion injury in rats

Myocardial ischemia–reperfusion (I/R) represents a clinically relevant problem associated with thrombolysis, angioplasty, and coronary bypass surgery. Radical oxygen species generated during early reperfusion are the primary activator of mitochondrial permeability transition pore (MPTP) opening which finally results in cardiomyocyte death. Nigella sativa (NS) has been shown to have antioxidant properties. The present study aimed to determine whether supplementation with NS can provide sufficient protection for the myocardium against I/R insult and any possible role on mitochondrial MPTP. Adult male Wistar rats were allocated into two groups: control group and NS-treated group receiving NS (800 mg/kg) orally for 12 weeks. Rats' isolated hearts were perfused in Langendorff preparation to determine the baseline heart beating rate, developed peak tension, time to peak tension, rate of tension development, half relaxation time, and myocardial flow rate. Ischemia was then induced by stopping the perfusion fluid for 30 min, followed by 30 min of reperfusion and recording post I/R cardiac functions. Hearts were then used for assessment of malondialdehyde (MDA) and nicotinamide adenine dinucleotide (NAD⁺), since the hydrolysis of mitochondrial NAD⁺ directly reflects MPTP opening in situ, and for histological examination. The NS-treated group showed enhanced post I/R contractile and vascular recovery, which was accompanied by elevated NAD⁺ and decreased MDA compared to the control group. Histological examination showed marked improvement of cardiac musculature compared to the control group. In conclusion, N. sativa afforded substantial recovery of post I/R cardiac functions probably via inhibition of MPTP opening.     

Protocol for Callus Induction and Somatic Embryogenesis in Moso Bamboo

Moso bamboo [Phyllostachys heterocycla var. pubescens (Mazel ex J. Houz.) Ohwi] is one of the most important forest crops in China and the rest of Asia. Although many sympodial bamboo tissue culture protocols have been established, there is no protocol available for plantlet regeneration as indicated by callus induction for monopodial bamboos, such as Moso bamboo. In the present report, embryogenic callus induction, embryoid development, and germination were established for Moso bamboo from zygotic seed embryos. Callus was initiated from zygotic embryos after 10–20 d culture on MS media supplemented with 4.0 mg/L 2, 4-D and 0.1 mg/L zeatin (ZT). About 50% of the explants produced calli, and nearly 15% of the calli were found to be embryogenic in nature. These embryogenic calli can be subcultured for proliferation in the Murashige and Skoog media (MS) supplemented with 0.5–2.0 mg/L 2, 4-D. These calli were found to have maintained their capacity for regeneration even after one year of subculture. The viable somatic embryoids regenerated in medium containing 5.0–7.0 mg/L ZT. Nearly 5% of the calli were found capable of regenerating into plantlets directly in MS medium containing 5.0–7.0 mg/L ZT. Root growth was more pronounced when the plantlets were transferred to medium containing 2.0 mg/L NAA. After 30 days of subculture, the plantlets were transferred to a greenhouse.     

蓝莓离体叶片胚状体高效发生及其组织学观察

以高灌蓝莓试管苗叶片为外植体、以改良WPM为基本培养基,研究了外源激素TDZ、ZT及其组合对离体叶片胚状体发生的影响,同时也探讨了蔗糖浓度、水解酪蛋白、椰汁等对胚状体发生、丛芽形成的影响.结果表明:不同浓度的外源激素TDZ、ZT及其组合对胚状体的发生频率、丛芽的形成和生长起重要作用;蔗糖浓度对胚状体的发生及丛芽的生长影响较大,而添加有机质则对胚状体的发生及丛芽的生长没有明显影响.适合高灌蓝莓叶片胚状体发生及成苗的培养基为WPM TDZ 0.04 mg/L ZT 0.25～2.0 mg/L 蔗糖20～40 g/L,而培养基WPM ZT 0.5～1.0 mg/L 蔗糖20 g/L适合于丛芽继代生长.组织学观察表明,蓝莓叶片胚状体发生主要起源于叶上表皮细胞和部分叶肉细胞,可能为多细胞起源,历经多细胞原胚、原球胚、梨形胚、心形胚、子叶胚等发育阶段,并能直接发育成苗.     

水蕨愈伤胚胎发生的代谢组学研究

体细胞胚胎发生是植物繁殖的一种方式,而离体的愈伤胚胎发生既可以进行植株再生、又可以用于遗传转化。该研究采用非靶向代谢组学,对蕨类植物水蕨分别处于增殖期和分化期的愈伤进行代谢物差异分析,以探讨愈伤胚胎发生的代谢机制,为愈伤胚胎发生的代谢组学提供资料。结果发现:(1)水蕨增殖期与分化期的愈伤具有相同的代谢物,且其中一些为疾病药物如放线菌酮和三七素。(2)各代谢物的含量高低不一,但基本上在增殖期和分化期之间无差异。(3)增殖期积累的代谢物预测与ABC通道蛋白途径有关,而分化期积累的代谢物可能与黄酮合成途径有关。研究推测,水蕨愈伤增殖期到分化期的转变是代谢物的量变过程,而非质变;植物愈伤胚胎发生的代谢组可能具有物种特异性。     

Anti-inflammatory Effect of Seeds and Callus of Nigella sativa L. Extracts on Mix Glial Cells with Regard to Their Thymoquinone Content

Anti-inflammatory effect of the alcoholic extracts of N. sativa seeds and its callus on mix glial cells of rat with regard to their thymoquinone (TQ) content was investigated. Callus induction was achieved for explants of young leaf, stem, petiole, and root of N. sativa on solid Murashige and Skoog (MS) medium containing 2,4-D (1 mg/l) and kinetin (2.15 mg/l). TQ content of the alcoholic extracts was measured by HPLC. Total phenols were determined using Folin–Ciocalteu method and antioxidant power was estimated using FRAP tests. The mix glial cells, inflamed by lipopolysaccharide, were subjected to anti-inflammatory studies in the presence of various amounts of TQ and the alcoholic extracts. Viability of the cells and nitric oxide production were measured by MTT and Griess reagent, respectively. The leaf callus obtained the highest growth rate (115.4 mg/day) on MS medium containing 2,4-D (0.22 mg/l) and kinetin (2.15 mg/l). Analyses confirmed that TQ content of the callus of leaf was 12 times higher than that measured in the seeds extract. However, it decreased as the calli aged. Decrease in the TQ content of the callus was accompanied with an increase in its phenolic content and antioxidant ability. Studies on the inflamed rat mix glial cells revealed significant reduction in the nitric oxide production in the presence of 0.2 to 1.6 mg/ml of callus extract and 1.25 to 20 μl/ml of the seed extracts. However, the extent of the effects is modified assumingly due to the presence of the other existing substances in the extracts.     

芍药花药愈伤组织诱导及体细胞胚发生

以芍药(Paeonia lactiflora)品种粉玉奴花药为外植体,研究不同浓度2,4-D对愈伤组织诱导、体胚发生及植株再生的影响,采用组织细胞学方法观察愈伤组织以及体细胞胚发育过程,采用根尖染色体法鉴定再生植株倍性。结果表明,芍药花药愈伤组织诱导的最适培养基为MS+1 mg·L–1 2,4-D+1 mg·L–1 N...     

Biochemical effects of Nigella sativa L seeds in diabetic rats

Oral administration of ethanol extract of N. sativa seeds (300 mg/kg body weight/day) to streptozotocin induced diabetic rats for 30 days significantly reduced the elevated levels of blood glucose, lipids, plasma insulin and improved altered levels of lipid peroxidation products (TBARS and hydroperoxides) and antioxidant enzymes like catalase, superoxide dismutase, reduced glutathione and glutathione peroxidase in liver and kidney. The results confirm the antidiabetic activity of N. sativa seeds extract and suggest that because of its antioxidant effects its administration may be useful in controlling the diabetic complications in experimental diabetic rats.     

Morphogenesis in Tissue Cultures of Different Organs of Nigella sativa

Calli were isolated from root, stem and leaf segments of Nigella sativa (Fam. Ranunculaceae) on White's medium containing napthaleneacetic acid and coconut milk. From all the three types of calli, roots were differentiated. Shoot development occurred in stem and leaf calli only after the omission of first coconut milk and then of auxin from the basal medium and also in IAA (2.0 mg/ml) and coconut milk (15 % v/v) in the medium. Frequency of organ formation and the maintenance of this capacity depend upon the nature as well as the age of the callus tissue.     

芍药愈伤组织中体细胞胚发育过程的组织细胞学观察

以芍药（Paeonia lactiflora Pall.）3个芍药品种的茎段、叶片、叶柄为外植体,诱导体细胞胚发生,并采用石蜡切片法对该发育过程进行组织细胞学观察。结果表明：‘Going Bananas’的茎段愈伤诱导率达100%,增殖率在4.0以上,表现最佳;非胚性愈伤组织最佳诱导、增殖培养基为WPM+IAA 1.0mg·L^-1+6-BA 1.0mg·L^-1+NAA 0.5mg·L^-1+TDZ 0.5mg·L^-1+CH 0.625g·L^-1;愈伤组织转入到1/2 MS（Ca²⁺加倍）+2,4-D 2.0mg·L^-1+ABA 0.5mg·L^-1或ZT 1.0mg·L^-1的培养基中,连续暗培养90后得到胚性愈伤;之后转入到成熟培养基1/2MS（Ca²⁺加倍）+6-BA 1.0mg·L^-1+NAA 0.2mg·L^-1中,见光培养60d,逐渐发育至球形胚和心形胚。在石蜡切片观察中,芍药的体细胞胚起源方式包括外起源和内起源两个途径。在外起源方式中包括单个表层细胞的外起源和多个表层下细胞的共同起源。3种方式的区别主要在于起始的位置和起始细胞数量,后期均形成原胚结构。胚性细胞的分裂方式为对称分裂,未发现不对称分裂细胞的存在。     

"灵发素"直接诱导马铃薯愈伤组织分化试管薯

1 植物名称马铃薯(Solanum tuberosum)中薯2号. 　　2 材料类别无菌试管苗. 　　3 培养条件愈伤组织诱导培养基:(1)MS 2,4-D 2 mg*L-1(单位下同).诱导块茎培养基:(2)MS 2,4-D 2 “灵发素“[1,2] 7.以上培养基均加0.7%琼脂、3%蔗糖,pH 5.8.放置于实验室内,没有设置特定温度与光照条件. 　　……     

NAA-Induced Organogenesis and Embryogenesis in Hypocotyl Callus of Solarium melongena L.

Hypocotyl explants of S. melongena showed three types of regenerationthrough callus formation depending on the concentration of NAAin the medium. At 0.8 mg l^–1, only callus was produced.Lower concentrations resulted in callus, adventitious roots(optimum, 0.016 mg 1^–1 NAA), and adventitious shoots (noNAA). Roots and shoots developed during the early stages ofculture. Higher concentrations of NAA depressed callus growthand stimulated embryoid formation (optimum 8.0 mg 1^–1NAA), Embryoids were identifiable after about 6 weeks as greenspots on the surface of callus: Addition of 6-BA enhanced shootproduction but inhibited both root and embryoid production.Whole plants were obtained from embryogenic callus after transferto NAA free medium. Genotypic differences in response were observed. In general,the potential for embryogenesis was independent of or inverselyrelated to the potential for organogenesis.     

Protective Effect of Nigella sativa and Nigella damascena Fixed Oils Against Aflatoxin Induced Mutagenicity in the Classical and Modified Ames Test

The antioxidant and mutagenic/antimutagenic activities of the fixed oils from Nigella sativa (NSO) and Nigella damascena (NDO) seeds, obtained by cold press-extraction from the cultivar samples, were comparatively investigated for the first time. The antimutagenicity test was carried out using classical and modified Ames tests. The fatty acid composition of the fixed oils was characterized by gas chromatography–mass spectrometry (GC-MS) while the quantification of thymoquinone in the fixed oils was determined by UPC². The main components of the NSO and NDO were found to be linoleic acid, oleic acid, and palmitic acid. The results of the Ames test confirmed the safety of NSO and NDO from the viewpoint of mutagenicity. The results of the three antioxidant test methods were correlated with each other, indicating NDO as having a superior antioxidant activity, when compared to the NSO. Both NSO and NDO exhibited a significant protective effect against the mutagenicity induced by aflatoxin B1 in Salmonella typhimurium TA98 and TA100 strains. When microsomal metabolism was terminated after metabolic activation of the mycotoxin, a significant increase in antimutagenic activity was observed, suggesting that the degradation of aflatoxin B1 epoxides by these oils may be a possible antimutagenic mechanism. It is worthy to note that this is the first study to assess the mutagenicity of NSO and NDO according to the OECD 471 guideline and to investigate antimutagenicity of NDO in comparison to NSO against aflatoxin.