苦荞胚性愈伤组织诱导与植株再生研究

以苦荞子叶和下胚轴为外植体,进行了不同浓度激素组合的MS和SH固体培养基对胚性愈伤组织诱导及植株再生的研究。结果发现,MS培养基比SH培养基更有利于胚性愈伤组织诱导;2,4-D是诱导愈伤组织的有效激素,KT能有效促进胚状体的形成;下胚轴和子叶都能有效诱导出胚性愈伤组织和再生植株。下胚轴在MS 1.5mg·L-12,4-D 1.5mg·L-1BA培养基,子叶在MS 2mg·L-12,4-D 0.5~1.5mg·L-1BA上能高效诱导出愈伤组织;愈伤组织在MS 2mg·L-12,4-D 0.1mg·L-1KT培养基中继代,能有效诱导胚性愈伤组织;来自下胚轴的胚性愈伤组织在1/2MS 2.0mg·L-1BA 0.5mg·L-1KT 0.1mg·L-1NAA培养基上能够高频再生出芽,来自子叶的胚性愈伤组织在1/2MS 1.0mg·L-1BA 0.1mg·L-1KT 0.1mg·L-1NAA培养基上芽诱导率较高;MS 1mg·L-1NAA是适宜的再生苗生根培养基。     

外源激素对杜衡叶柄愈伤组织诱导与分化的影响

以杜衡(Asarum forbesii Maxim.)叶柄为外植体,采用L9(34)正交实验设计分别研究了培养基中外源激素的种类及质量浓度对杜衡叶柄愈伤组织诱导和分化的影响,并据此筛选出适宜的诱导及分化培养基.结果表明:在杜衡叶柄愈伤组织的诱导过程中,培养基中细胞分裂素的种类(1.00 mg·L-16-BA、1.00 mg·L-1KT和1.00 mg·L-1ZT)和NAA质量浓度(0.00、0.10和0.30 mg·L-1)的影响效应均不显著,而2,4-D质量浓度(0.10、0.50和1.00 mg·L-1)则有显著影响(P<0.05);在愈伤组织的分化培养过程中,NAA(0.10、0.30和0.50 mg·L-1)的影响效应大于6-BA(1.00、3.00和5.00 mg·L-1)和IBA(0.01、0.05和0.10 mg·L-1).综合比较结果显示,适宜于杜衡叶柄愈伤组织诱导的培养基为添加1.00 mg·L-16-BA、0.30 mg·L-1NAA和1.00 mg·L-12,4-D的MS培养基(含6.5 g·L-1琼脂和30 g·L-1蔗糖,pH 5.8～pH 6.0),在此培养基上愈伤组织诱导率达到83.33%,且愈伤组织生长速度快、颗粒紧密;适宜于杜衡愈伤组织分化和不定芽增殖的培养基为添加3.00 mg·L-16-BA、0.10 mg·L-1IBA和0.30 mg·L-1NAA的MS培养基(含6.5 g·L-1琼脂和30 g·L-1蔗糖,pH 5.8～pH 6.0),在此培养基上愈伤组织的分化率最高(达到53.33%),增殖系数也最高(3.13).     

海滨锦葵胚轴愈伤组织诱导及植株再生

以海滨锦葵(Kosteletzkya virginica)胚轴为外植体, 在9种不同激素配比的培养基上进行愈伤组织诱导、继代培养、不定芽分化及生根培养, 确定了植株再生的最适培养条件: (1)愈伤组织诱导最适培养基为MS + IAA 1.0 mg.L-1 + KT 0.3 mg.L-1 + sucrose 30 g.L-1 + agar 8 g.L-1, 愈伤组织诱导率为93.94%; (2)不定芽诱导最适培养基为MS + IAA 0.1 mg.L-1 + ZT 0.5 mg.L-1 + sucrose 30 g.L-1 + agar 8 g.L-1, 不定芽诱导率为65.83%; (3)生根最适培养基为MS + sucrose 30 g.L-1 +agar 8 g.L-1, 生根率为96.67%。炼苗移栽后, 成活率可达85%。     

甜高粱再生体系的建立

利用甜高粱成熟种子和成熟胚作为外植体, 通过愈伤组织再生途径建立了植株再生体系。结果表明, 成熟种子在添加1.38 g.L-1脯氨酸、500 mg.L-1水解酪蛋白和3.0 mg.L-1 2, 4-D的MS培养基上可以较快地诱导出生长状态良好的愈伤组织;成熟胚在添加1.38 g.L-1脯氨酸、500 mg.L-1水解酪蛋白、2.5 mg.L-1 2, 4-D和0.1 mg.L-1KT的MS培养基上也能诱导出生长良好的愈伤组织。将愈伤组织转移到MS+1 mg.L-1 IAA + 0.5 mg.L-1 6-BA培养基上诱导芽, 然后再转移到MS+3 mg.L- 1IBA培养基上生根后, 可发育成为完整的植株。     

一种改进的水稻成熟胚愈伤组织高效基因转化系统

以成熟胚愈伤组织为材料的农杆菌介导水稻转化法虽已建立, 但转化频率仍有待提高。本文以粳稻(Oryz a sativa)品种(中花10号和中花11号)的成熟胚诱导的愈伤组织为受体材料, 对组织培养体系及影响遗传转化的因素进行优化, 建立了一套改进的农杆菌介导的水稻高效遗传转化系统。农杆菌菌株为EHA105, 质粒载体是pUN1301/ OsRAA1, 其中含有标记基因GUS 和筛选基因HPT。愈伤组织诱导培养基为NBD2 (NB+2 mg·L-12,4-D), 继代培养基为NBD0.5, 预分化与分化培养基为RE1 (MS+1 mg·L-16-BA + 0.25 mg·L-1 NAA + 0.5 mg·L-1 KT + 0.2 mg·L-1 ZT)和RE2 (MS+ 1 mg·L-1 6-BA + 0.5 mg·L-1 NAA+ 0.5 mg·L-1 KT + 0.2 mg·L-1 ZT)。另外, 还分析了影响T-DNA转移的多种因素, 如外植体种类、愈伤组织预培养基和愈伤组织继代次数等。采用优化的转化程序, 水稻愈伤组织转化率和植株转化率可达70%以上。     

甜高粱再生体系的建立

利用甜高粱成熟种子和成熟胚作为外植体,通过愈伤组织再生途径建立了植株再生体系。结果表明,成熟种子在添加1.38 g·L^-1脯氨酸、500 mg·L^-1水解酪蛋白和3.0 mg·L^-1 2,4-D的MS培养基上可以较快地诱导出生长状态良好的愈伤组织;成熟胚在添加1.38 g·L^-1脯氨酸、500mg·L^-1水解酪蛋白、2.5 mg·L^-1 2,4-D和0.1 mg·L^-1KT的MS培养基上也能诱导出生长良好的愈伤组织。将愈伤组织转移到MS＋1 mg.L^-1 IAA＋0.5 mg·L^-1 6-BA培养基上诱导芽,然后再转移到MS＋3 mg·L^-1IBA培养基上生根后,可发育成为完整的植株。     

一种改进的水稻成熟胚愈伤组织高效基因转化系统

以成熟胚愈伤组织为材料的农杆菌介导水稻转化法虽已建立,但转化频率仍有待提高.本文以粳稻(Oryza sativa)品种(中花10号和中花11号)的成熟胚诱导的愈伤组织为受体材料,对组织培养体系及影响遗传转化的因素进行优化,建立了一套改进的农杆菌介导的水稻高效遗传转化系统.农杆菌菌株为EHA105,质粒载体是pUN1301/OsRAA1,其中含有标记基因GUS和筛选基因HPT.愈伤组织诱导培养基为NBD2(NB 2 mg·L-12,4-D),继代培养基为NBD0.5,预分化与分化培养基为RE1(MS 1 mg·L-16-BA 0.25 mg·L-1NAA 0.5 mg·L-1KT 0.2 mg·L-1ZT)和RE2(MS 1 mg·L-16-BA 0.5 mg·L-1NAA 0.5 mg·L-1KT 0.2 mg·L-1ZT).另外,还分析了影响T-DNA转移的多种因素,如外植体种类、愈伤组织预培养基和愈伤组织继代次数等.采用优化的转化程序,水稻愈伤组织转化率和植株转化率可达70%以上.     

芒属植物荻愈伤组织的诱导与植株再生

利用资源植物荻(Miscanthus sacchariflorus)的幼穗作为外植体,通过愈伤组织途径建立了植株再生体系.结果表明:在附加600 mg·L-1脯氨酸、500 mg·L-1水解酪蛋白、2.0mg ·L-12,4-D和0.1mg·L-16-BA的MS培养基上能高效诱导出生长良好的愈伤组织;将其转移到MS+1.5 mg·L-1 6-BA培养基上诱导产生芽,再转移到1/2 MS+0.25 mg·L-1 NAA+0.25 mg·L-1 IAA培养基上生根后,可发育为生长健壮的植株.     

海南粗榧愈伤组织的诱导和培养

海南粗榧的嫩茎和嫩叶以0.1%升汞消毒12 min的效果最好.外植体在MS 0.1 mg·L.6-BA l mg·L-1NAA 2mg·L-12,4-D 3.0%蔗糖 0.6%卡拉胶(pH 5.8)培养基中能顺利诱导出愈伤组织.液体悬浮和暗培养均有利于愈伤组织的增殖.愈伤组织增殖的最佳培养基为MS 0.1 mg·L-1KT 3 mg·L-1NAA 3.0%蔗糖(pH 5.8).     

沙拐枣的组织培养

1 植物名称沙拐枣(Calligonum mongolicwn Turvz.). 2 材料类别由种子获得的小苗茎段. 3 培养条件愈伤组织诱导培养基:(1)MS+6-BA0.05 mg·L-1(单位下同)+IAA 0.01;愈伤组织增殖培养基:(2)MS+6-BA 0.4;愈伤组织分化培养基:(3)MS+6-BA 0.4+2,4-D 0.5;继代培养基:(4)MS+IAA 0.1+6-BA 0.05;生根培养基:(5)MS+IAA 0.1.     

Reduction of auxin transport capacity with age and internal water deficits in cotton petioles

Auxin transport was examined in leaf petioles taken from the upper, middle, and lower leaf canopy of large cotton plants. The ability of petioles to transport auxin decreased with age (position) of the leaves. Plant water deficit reduced transport regardless of age. These correlations support the view that reduced transport capacity of petioles plays a significant role in the induction of abscission of lower or older leaves during water deficits.     

Stimulation of ethylene evolution and abscission in cotton by 2-chloroethanephosphonic Acid

Ethrel, a mixture of 2-chloroethanephosphonic acid and its ethyl ester, hastens abscission of leaves, debladed petioles, and flower buds of cotton plants (Gossypium hirsutum, L.). Both young and old leaves abscissed while still green. Application of Ethrel stimulated evolution of ethylene, and this response preceded abscission. Air concentrations of ethylene around enclosed, treated-plants were adequate to produce abscission in plants. Non-treated plants defoliated when enclosed with plants sprayed with Ethrel. The stimulation of abscission of explant petioles by Ethrel was reversed by naphthalene acetic acid. The stimulation of abscission by Ethrel was concluded to be mediated by ethylene.     

Differential expression of a chlorophyll a/b binding protein gene and a proline rich protein gene in juvenile and mature phase English ivy (Hedera helix)

Two cDNA clones representing mRNAs which are differentially expressed during in vitro culture of juvenile and mature leaf petioles of English ivy ( Hedera helix L.) were isolated by differential screening. The mRNA represented by clone HW101 is expressed at a higher level in untreated juvenile than in untreated mature in-vitro-cultured petioles. Treatment of petioles with α-naphthaleneacetic acid (NAA) at the initiation of culture decreases HW101 mRNA levels in juvenile but not mature, petioles. In intact plants. HW101 mRNA is expressed at a higher level in juvenile laminae, petioles and stems than in identical tissues of mature plants. DNA sequence analysis indicates that HW1O1 cDNA is significantly similar to a light harvesting chlorophyll a/b binding protein gene ( Lhcb ) of pea. The gene represented by the second clone. HW103, is expressed at a higher level in mature than in juvenile in-vitro-cultured petisoles. Treatment of petioles with NAA at the initiation of culture decreases HW103 mRNA levels in chronologically young mature but not older mature and juvenile petioles. However, expression of the HW103 gene is not detectable in petioles, or in any other vegetative organ tested, immediately after excision. It is, however, expressed in developing seeds. In otherwise intact plants, the HW103 gene is expressed in wounded petioles of mature plants 5 days after wounding but not in wounded petioles of juvenile plants. It is also expressed at a higher level in wounded stems of mature plants than in those of juvenile plants. However, it is not expressed in wounded lamina of either juvenile or mature plants. DNA sequence analysis indicates that HW103 cDNA is similar to a cell wall proline rich protein (PRP) gene of soybean. This is the first report of differential expression of a PRP gene in tissues from juvenile and mature plants. Southern blot analysis of nuclear DNA of H. helix shows that both HW101 and HW103 are members of small gene families.     

Cellulolytic Activity of Botrytis cinerea in vitro and in vivo

Experiments were carried out to determine whether stepwise breakdown of native cellulose is carried out by B. cinereain vitro and in vivo. Protein fractions were obtained from ungerminated conidia, from culture filtrates 24, 48 and 96 h after inoculation with conidia, and from culture filtrates of 12 day-old cultures growing on cotton wool as the carbon source. In addition, petioles and fruits of tomato plants were inoculated and the protein fraction of the colonized tissues were tested. Using filter paper, carboxymethylcellulose and cellobiose as substrates, all fraction showed c₁, glucanase and cellobiase activity respectively.     

Effects of fusicoccin on intact cotton plants (Gossypium hirsutum L.)

Fusicoccin (FC) was applied as a spray to shoots of intact field- and glasshouse-grown cotton plants. Distortions of shoot morphology resulted. Stems and petioles of FC-treated plants were irregular in diameter and twisted, whereas leaf laminae were curled and crinkled. Shoot elongation was inhibited by FC; the effect was dependent upon the concentration and timing of the applications.Abbreviation FC fusicoccin     

Absorption and translocation of sodium in beans and cotton

At the end of a 4 hour absorption period approximately 95% of the sodium absorbed by bean plants was retained in the secondary roots. The sodium translocated to the shoot was retained in the stem.

2,4-Dinitrophenol decreased the amount retained in the secondary roots of bean plants and increased the amount translocated to the shoot. The stem retained most of the translocated sodium.

Bean plants without roots absorbed considerably more sodium than plants with roots and translocated a greater proportion of the sodium to the petioles and blades. 2,4-Dinitrophenol reduced the amount of sodium in the stem and petioles and increased the amount in the blades.

2,4-Dinitrophenol reduced the amount of sodium retained by the secondary roots of cotton plants but did not appreciably affect the amounts translocated to the shoot.

     

In Vitro Shoot Regeneration from Callus, Leaf Axils and Petioles of Sugar Beet (Beta vulgaris L.)

Procedures are described for producing axillary shoots fromseedling apices and adventitious shoots from petioles and leaf-derivedcallus of sugar beet cultivars. The rate of adventitious shootregeneration from petioles was influenced by temperature, BAPconcentration of the medium, and the time in culture of theseedling apices from which the petioles were excised. Petiolesectioning confirmed that adventitious shoots originated inthe sub-epidermal parenchyma. Two distinct types of callus wereproduced from leaf explants, but only white friable callus wascapable of shoot development. This callus developed from browntissue and was composed of thin-walled cells with dense cytoplasmand prominent nuclei. Green compact callus with thick-walledlignified cells developed from green tissue, but did not produceshoots. Successful seed sterilization and shoot regenerationfrom petiole explants and callus was cultivar-dependent. Adventitiousshoots were rooted and successfully transplanted to pottingcompost under glasshouse conditions. Key words: Adventitious shoots, axillary shoots, callus, sugar beet (Beta vulgaris L.)     

The Commelina Yellow Mottle Virus Promoter Is a Strong Promoter in Vascular Tissue of Transgenic Gossypium hirsutum Plants

Explants of cotton (Gossypium hirsutum L. cv. Jingmian 7) were transformed with Agrobacterium tumefaciens (Smith et Townsend ) Conn LBA4404 harboring an expression cassette composed of CoYMV (Commelina Yellow Mottle Virus) promoter-gus-nos terminator on the plant expression vector pBcopd2. Transgenic plants were regenerated and selected on a medium containing kanamycin. GUS (β-glucuronidase) activity assays and Southern blot analysis confirmed that the chimerical gus gene was integrated into and expressed in the regenerated cotton plants. Plant expression vector pBI121 was also transferred into the same cotton variety and the regenerated transgenic plants were used as a positive control in GUS activity analysis. Evidences from histochemical analysis of GUS activity demonstrated that under the control of a 597 bp CoYMV promoter the gus gene was highly expressed in the vascular tissues of leaves, petioles, stems, roots, hypocotyls, bracteal leaves and most of the flower parts while GUS activity could not be detected in stigma, anther sac and developing cotton fibers of the transgenic cotton plants. GUS specific activity in various organs and tissues from transgenic cotton lines was determined and the results indicated that the CoYMV promoter-gus activities were at the same level or higher than that of CaMV 35S promoter-gus in leaf veins and roots where the vascular tissues occupy a relatively larger part of the organs, but in other organs like leaves, cotyledons and hypocotyls where the vascular tissues occupy a smaller part of the organs the CoYMV promoter-gus activity was only 1/3-1/5 of the CaMV 35S promoter-gus activity. The GUS activity ratio between veins and leaves was averaged 0.5 for 35S-GUS plants and about 2.0 for CoYMV promoter-gus transgenic plants. These results further demonstrated the vascular specific property of the promoter in transgenic cotton plants. An increasing trend of GUS activity in leaf vascular tissues of transgenic cotton plants developing from young to older was observed.     

Nitrogen fertilization rate affects feeding, larval performance, and oviposition preference of the beet armyworm, Spodoptera exigua, on cotton

Nitrogen (N) is one of the most critical chemical elements for plant and animal growth, exerting a variety of bottom‐up effects. Development and oviposition of the beet armyworm, Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae), were studied in relation to varying N fertilization levels (42, 112, 196, and 280 p.p.m.) in cotton [Gossypium hirsutum L. (Malvaceae)]. Low N fertilization of cotton plants led to reduced plant biomass and a lower percentage of N in leaf blades and in leaf petioles. Development of S. exigua larvae fed with plants with reduced N applications (42 and 112 p.p.m.) was prolonged relative to treatments receiving higher N fertilization. Almost all larvae reared on artificial diets underwent only five instars before pupation. However, most larvae reared on cotton plants, irrespective of N levels, experienced a supernumerary sixth larval instar. Furthermore, significantly more larvae reared on lower N cotton plants underwent supernumerary development compared to larvae reared on higher N cotton plants. Life‐time feeding damage per larva ranged from 55 to 65 cm², depending on the nutritional quality of the food plant, although the differences were not statistically significant. Larvae distinguished between cotton plants with various nutritional qualities and fed preferentially on higher N plants. Female moth oviposition choice was also affected by host plant nutritional quality: cotton plants with higher N levels were preferentially chosen by S. exigua females for oviposition. The mechanisms of these effects are unclear, but they can have important implications for population dynamics and pest status of beet armyworms in the field.     

PHYTOTOXIC EXUDATES FROM VELVETLEAF (ABUTILON THEOPHRASTI) GLANDULAR TRICHOMES

Nonvolatile exudates from velvetleaf glandular trichomes inhibited root and shoot growth of several weed and crop species in petri plate bioassays, but had no effect on seed germination per se. The exudate was efficiently collected by wiping both the stems and petioles with cotton swabs or by leaching with water, but was absent on the leaves of velvetleaf plants. Cress (Lepidium sativum L.) was the most sensitive indicator species. Four types of trichomes appeared on the stem surface as revealed by scanning electron microscopy. Water soluble globules on the apices of 12- to 15-celled glandular trichomes recurred and demonstrated their original potency within eight days after removal with cotton swabs. Both the quantity and phytotoxicity of the exudates from velvetleaf plants cultured under varying environmental conditions were determined. While total exudate production was not affected at 16, 24, or 36 C, the exudates from plants cultured at 24 and 36 C were about twice as toxic as the exudate collected from plants grown at the lower temperature. Water stress decreased the amount of exudate collected, but the phytotoxic activity was increased by approximately the same factor.