Growth and physiological responses of cotton (Gossypium hirsutum L.) to elevated carbon dioxide and ultraviolet-B radiation under controlled environmental conditions

Better understanding of crop responses to projected changes in climate is an important requirement. An experiment was conducted in sunlit, controlled environment chambers known as soil–plant–atmosphere–research units to determine the interactive effects of atmospheric carbon dioxide concentration [CO₂] and ultraviolet‐B (UV‐B) radiation on cotton (Gossypium hirsutum L.) growth, development and leaf photosynthetic characteristics. Six treatments were used, comprising two levels of [CO₂] (360 and 720 µmol mol^?1) and three levels of 0 (control), 7.7 and 15.1 kJ m^?2 d^?1 biologically effective UV‐B radiations within each CO₂ level. Treatments were imposed for 66 d from emergence until 3 weeks after the first flower stage. Plants grown in elevated [CO₂] had greater leaf area and higher leaf photosynthesis, non‐structural carbohydrates, and total biomass than plants in ambient [CO₂]. Neither dry matter partitioning among plant organs nor pigment concentrations was affected by elevated [CO₂]. On the other hand, high UV‐B (15.1 kJ m^?2 d^?1) radiation treatment altered growth resulting in shorter stem and branch lengths and smaller leaf area. Shorter plants at high UV‐B radiation were related to internode lengths rather than the number of mainstem nodes. Fruit dry matter accumulation was most sensitive to UV‐B radiation due to fruit abscission. Even under 7.7 kJ m^?2 d^?1 of UV‐B radiation, fruit dry weight was significantly lower than the control although total biomass and leaf photosynthesis did not differ from the control. The UV‐B radiation of 15.1 kJ m^?2 d^?1 reduced both total (43%) and fruit (88%) dry weights due to smaller leaf area and lower leaf net photosynthesis. Elevated [CO₂] did not ameliorate the adverse effects of UV‐B radiation on cotton growth and physiology, particularly the boll retention under UV‐B stress.     

Rapid in-vitro plant regeneration of cotton (Gossypium hirsutum L.)

A rapid, clonal propagation procedure has been developed to regenerate mature cotton (Gossypium hirsutum L.) plants from pre-existing meristems that were excised from in-vitro-grown tissues. This plant regeneration procedure was applicable to diverse cotton germplasms and required specific concentrations of 6-benzylaminopurine (BA) depending on the origin of the meristems. All shoots regenerated directly without a callus phase. Screening BA concentrations (0.0–10.0 μm) demonstrated that shoot meristems (apices), secondary leaf nodes, primary leaf nodes, and cotyledonary nodes derived from in-vitro-grown 28-day-old seedlings (Paymaster HS26) varied in their ability to form elongated shoots depending on the level of BA. Indicative of a germplasm-independent procedure, a BA concentration screen (0.0, 0.3, 1.0 μm) demonstrated that explants with pre-existing meristems, excised from diverse germlines, were also able to form elongated shoots at 0.3 μm BA. In most cases, elongated shoots derived from this procedure were rooted by a two-step process: an in-vitro maturation step (Murashige and Skoog medium-activated charcoal) followed by planting into soil after basal application of Rootone. This BA plant regeneration procedure was rapid, reproducible, and highly efficient for Stoneville 7A, Paymaster HS26, and other high-fiber-yielding germlines. Regenerated plants were phenotypically normal and all of the mature plants regenerated to date have initiated flowers and set viable R₁ seeds. Received: 15 March 1997 / Revision received: 28 August 1997 / Accepted: 5 September 1997     

Characterization of somatic embryogenesis and plant regeneration in cotton (Gossypium hirsutum L.)

Summary Seventeen cultivars of cotton (Gossypium hirsutum L.) were evaluated for callus initiation and maintenance using 3 initiation media and 3 maintenance media. After a series of transfers of a 3% glucose media, calli were placed on a 3% sucrose medium. After several weeks calli were observed for the presence of embryo-like structures. Cultivars Coker 201 and Coker 315 were identified as embryogenic. Embryogenic callus has since been routinely obtained within 6 weeks by initiating callus on glucose media for 3–4 weeks followed by transfer to sucrose media. Histological examination has shown that embryos are derived from isodiametric, densely cytoplasmic cells and follow predictable patterns of development. Upon maturity, transfer to auxin-free media with reduced sucrose levels results in embryo germination. Regenerated plants can be transferred to greenhouse within 90 days of callus initiation.The senior author is presently a Research Geneticist, USDA-ARS, and Assistant Professor Present address     

A genetic approach to in vitro regeneration of non-regenerating cotton (Gossypium hirsutum L.) cultivars

Selections were made among individual plants of Gossypium hirsutum cv `Coker 310' for high-frequency in vitro regeneration by somatic embryogenesis. After three generations of selection, a pure line for high-frequency somatic embryogenesis was selected and named Coker 310 FR (FR, fully regenerating). Coker 310 FR could be regenerated by following previously published protocols (see Materials and methods) and a modified protocol developed in this study that reduced the time necessary for in vitro regeneration. Coker 310 FR was crossed with individual plants of major cotton cultivars grown in India, namely `MCU 5', `MCU 7', `Khandwa 2', `Bikaneri Nerma', `F 846' that have been shown to be recalcitrant to in vitro regeneration, to evaluate the regeneration potential of F₁s. All the F₁s showed regeneration by somatic embryogenesis. However, the F₁ of G. barbadense×G. hirsutum Coker 310 FR did not regenerate. Received: 16 September 1997 / Revision received: 1 April 1998 / Accepted: 15 May 1998     

High-frequency stable transformation of cotton (Gossypium hirsutum L.) by particle bombardment of embryogenic cell suspension cultures

Stable transformation of cotton (Gossypium hirsutum L.) at a high frequency has been obtained by particle bombardment of embryogenic cell suspension cultures. Transient and stable expression of the β-glucuronidase (GUS) gene was monitored in cell suspension cultures. Transient expression, measured 48 h after bombardment, was abundant, and stable expression was observed in over 4% of the transiently expressing cells. The high efficiency of stable expression is due to the multiple bombardment of rapidly dividing cell suspension cultures and the selection for transformed cells by gradually increasing the concentrations of the antibiotic Geneticin (G418). Southern analysis indicated a minimum transgene copy number of one to four in randomly selected plants. Fertile plants were obtained from transformed cell cultures less than 3 months old. However, transgenic and control plants from cell cultures older than 6 months produced plants with abnormal morphology and a high degree of sterility. Received: 20 January 1999 / Revision received: 1 October 1999 / Accepted: 11 October 1999     

Comparative Development of Lint and Fuzz Using Different Cotton Fiber-specific Developmental Mutants in Gossypium hirsutum

A series of fiber-specific mutants, or germplasms, have been recently used in the study of fiber development. In thecurrent study, scanning electron microscopy (SEM) was used to investigate developmental differences in lint and fuzzinitiation in different genotypes (Gossypium hirsutum) of upland cotton.These fiber mutants included dominant nakedseed N1, recessive naked seed n2, Xuzhou-142 lintless-fuzzless (XZ142WX), Xinxiangxiaojilintless-fuzzless (XinWX),Xinxiangxiaojilinted-fuzzless (XinFLM), with TM-1, the cytogenetic and genetic experimental standard stock, as the control.Characteristics of fiber initiation were analyzed from -1 to 1 days post anthesis (dpa) and at 4 and 5 dpa for fuzz initiation.Our data suggested that lint initiation centered on day of anthesis (Odpa), and elongated significantly at 1dpa, while fuzzinitiation began at 4dpa, although the shape of fuzz protrusions differed from that of lint fibers. Fiber initiation occurred firston the ovule funicular crest. Compared to TM-1, there was a noted retardation in development and fiber protrusion in N1and XinFLM. Microscopy data also demonstrated that lintless-fuzzless mutants (XZ142WX and XinWX) developed irregularprotrusions during early developmental stages, which were unable to grow into fiber.     

Comparative Development of Lint and Fuzz Using Different Cotton Fiber-specific Developmental Mutants in Gossypium hirsutum

A series of fiber-specific mutants, or germplasms, have been recently used in the study of fiber development. In the current study, scanning electron microscopy （SEM） was used to investigate developmental differences in lint and fuzz initiation in different genotypes （Gossypium hirsutum） of upland cotton. These fiber mutants included dominant naked seed N1, recessive naked seed n2, Xuzhou-142 lintless-fuzzless （XZ142WX）, Xinxiangxiaojilintless-fuzzless （XinWX）, Xinxiangxiaojilinted-fuzzless （XinFLM）, with TM-1, the cytogenetic and genetic experimental standard stock, as the control. Characteristics of fiber initiation were analyzed from -1 to ＋1 days post anthesis （dpa） and at 4 and 5 dpa for fuzz initiation. Our data suggested that lint initiation centered on day of anthesis （0dpa）, and elongated significantly at 1dpa, while fuzz initiation began at 4dpa, although the shape of fuzz protrusions differed from that of lint fibers. Fiber initiation occurred first on the ovule funicular crest. Compared to TM-1, there was a noted retardation in development and fiber protrusion in N1 and XinFLM. Microscopy data also demonstrated that lintless-fuzzless mutants （XZ142WX and XinWX） developed irregular protrusions during early developmental stages, which were unable to grow into fiber.     

Response of the enzymes to nitrogen applications in cotton fiber (Gossypium hirsutum L.) and their relationships with fiber strength

To investigate the response of key enzymes to nitrogen (N) rates in cotton fiber and its relationship with fiber strength, experiments were conducted in 2005 and 2006 with cotton cultivars in Nanjing. Three N rates 0, 240 and 480 kgN/hm2, signifying optimum and excessive nitrogen application levels were applied.The activities and the gene expressions of the key enzymes were affected by N, and the characteristics of cellulose accumulation and fiber strength changed as the N rate varied. Beta-1,3-glucanase activity in cotton fiber declined from 9 DPA till boll opening, and the beta-1, 3-glucanase coding gene expression also followed a unimodal curve in 12—24 DPA. In 240 kgN/hm² condition, the characteristics of enzyme activity and gene expression manner for sucrose synthase and beta-1,3-glucanase in developing cotton fiber were more favorable for forming a longer and more steady cellulose accumulation process, and for high strength fiber development.     

Changes in the level of tubulin subunits during development of cotton (Gossypium hirsutum) fiber

The levels of tubulin protein in developing cotton ( Gossypium hirsutum L. cv. Stoneville 825) fibers were measured from 8 to 28 days post-anthesis using commercially available monoclonal antibodies against alpha- and beta-tubulin. As the monoclonal antibodies against alpha- and beta-tubulin were prepared from yeast tubulin and chick brain tubulin, respectively, indirect immunofluorescence microscopy was used to establish that the two monoclonal antibodies recognized microtubule structures in cotton fibers. Western blots of electrophoretically separated proteins in crude extracts of cotton roots and fibers showed that single polypeptides with the expected apparent molecular weight for tubulin subunits were recognized by the antisera. An enzyme-linked immunosorbent assay was used to quantify tubulin levels. From 10 to 20 days post-anthesis the level of tubulin protein increases approximately three-fold. After 20 days post-anthesis, the amount of tubulin relative to total fiber protein reaches a plateau or decreases slightly. The rapid rise in tubulin is correlated with the elongation of the fiber and an increase in cellulose synthesis.     

An RFLP linkage map of Upland cotton, Gossypium hirsutum L.

 Ninety-six F₂.F₃ bulked sampled plots of Upland cotton, Gossypium hirsutum L., from the cross of HS46×MARCABUCAG8US-1-88, were analyzed with 129 probe/enzyme combinations resulting in 138 RFLP loci. Of the 84 loci that segregated as co-dominant, 76 of these fit a normal 1 :  2 : 1 ratio (non-significant chi square at P=0.05). Of the 54 loci that segregated as dominant genotypes, 50 of these fit a normal 3: 1 ratio (non-significant chi square at P=0.05). These 138 loci were analyzed with the MAPMAKER∖ EXP program to determine linkage relationships among them. There were 120 loci arranged into 31 linkage groups. These covered 865 cM, or an estimated 18.6% of the cotton genome. The linkage groups ranged from two to ten loci each and ranged in size from 0.5 to 107 cM. Eighteen loci were not linked. Received: 31 March 1998 / Accepted: 29 April 1998     

棉花体细胞增殖和胚胎发生中的细胞程序性死亡

棉花组织培养中愈伤组织褐化可能与细胞程序性死亡(PCD)有关.对棉花组培中不同时期的愈伤组织DNA进行琼脂糖电泳,观察到:仅在第一次愈伤组织继代后10 d左右和愈伤组织第一次继代到分化培养基中培养10 d左右,愈伤组织的DNA产生了180 bp左右的片断或其整数倍片断大小的DNA带,呈DNA梯(DNA ladder)状电泳,说明在这两个时期PCD达到了高峰.在这两次PCD高峰后1周均出现愈伤组织大规模的褐化死亡.显微镜观察到第一次PCD发生高峰的愈伤组织存在许多管状、内含物少的细胞,这些细胞分布在愈伤组织的边缘和内部;第二次PCD发生高峰观察到体细胞胚胎分化、PCD细胞的木栓化和水渍化.对体细胞胚胎分化时期的原生质体进行荧光染色(DAPI),荧光显微镜下观察到发生细胞程序性死亡的细胞核浓缩、染色质凝聚、结块、边缘化和形成PCD小体.     

Indirect somatic embryogenesis and plant recovery from cotton (Gossypium hirsutum L.)

Summary We describe a tissue culture procedure for somatic embryogenesis and plantlet regeneration in cotton (Gossypium hirsutum L. cv. Coker 312). Callused explants or individual globular embryos were transferred to basal media to induce somatic embryogenesis. To determine characteristic early indicators of successful germination and conversion, we identified six types of embryos that developed on basal media. Two of the six embryo types, designated as tulip-shaped and trumpet-shaped, could undergo conversion in preliminary tests, whereas the others had little or no developmental potential. Several media treatments designed to enhance the maturation of globular somatic embryos failed to increase the fraction of embryos which matured to form recoverable types. In efforts to improve plantlet recovery, tulip-shaped embryos were used in limited trials to contrast the effects of chemical and physical desiccation treatments on germination and conversion. The selective use of tulip-shaped somatic embryos, coupled with partial desiccation, seems to have augmented plant recovery. Growth habit, flowering, seed set, and lint production of most of the regenerated plants were comparable to seed-derived plants grown under the same conditions. Partial research support was provided by state and federal funds appropriated to the Ohio Agricultural Research and Development Center, The Ohio State University.     

Somatic embryogenesis and plant regeneration in cotton (Gossypium hirsutum L.)

Tissue culture methods for improvement of cotton has lagged seriously compared to other major crops. A method for regeneration of cotton which includes a morphogenetically competent cell suspension was needed to facilitate selection of stress-resistant variants and gene manipulation. Preliminary screening of eight strains of Gossypium hirsutum L. for embryogenic potential resulted in the production of somatic embryos in all strains. Coker 312 was selected for use in the development of a model regeneration system for G. hirsutum. Calli were initiated from hypocotyl tissues of 3-day-old-seedlings. Globular embryos were present after six weeks in culture. Calli were subcultured to liquid suspension in growth regulator-free medium. After three to four weeks, suspensions were sieved to collect globular and heart stage embryos. Collected embryos developed further when plated onto semi-solid medium. To induce germination and plantlet growth, mature embryos were placed on sterile vermiculite saturated with medium. Upon development of roots and two true leaves, plantlets were potted in peat and sand, and hardened. Mature plants and progeny have been obtained with this procedure. A high percentage of infertile plants was observed among the regenerants.Abbreviations NAA 1 naphthaleneacetic acid - IAA indole-3-acetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - MS Murashige and Skoog - BA 6 benzylamino purine - 2i P N⁶-(²-isopentenyladenine     

陆地棉核不育株扦插繁殖与宿生保持及杂种优势利用

利用广西南宁冬季无霜冻的气候特点,对陆地棉(Gossypium hirsutum L.)洞A细胞核雄性不育扦插株与宿生株及其杂交一代进行比较研究.结果表明:(1)1 a生扦插株开花较早,茎粗、主茎节间长度、铃重、子指4个性状显著好于实生株,但扦插株的种子产量显著低于实生株,其原因是果枝扦插形成的僵苗占扦插株的23.04%,而且扦插株全为不育株,实生株中则有50%左右的可育株;(2)2 a生扦插株性状与实生株无显著差异;(3)由扦插繁殖的不育株和种子繁殖的不育株配制的杂交F1代的产量与纤维品质无显著差异.因此认为,扦插繁殖陆地棉核不育株用于多年杂交制种是可行的,选择营养枝扦插是陆地棉核不育株扦插繁殖需要注意的关键问题. 宿生株及其杂交一代进行比较研究.结果表明:(1)1 a生扦插株开花较早,茎粗、主茎节间长度、铃重、子指4个性状显著好于实生株,但扦插株的种子产量显著低于实生株,其原因是果枝扦插形成的僵苗占扦插株的23.04%,而且扦插株全为不育株,实生株中则有50%左右的可育株;(2)2 a生扦插株性状与实生株无显著差异;(3)由扦插繁殖的不育株和种子繁殖的不育株配制的杂交F1代的产量与纤维品质无显著差异.因此认 ,扦插繁殖陆地棉核不育株用于多年杂交制种是可行的,选择营养枝扦插是陆地棉核不育株扦插繁殖需     

The effects of UV-B radiation on epidermal anatomy in loblolly pine (Pinus taeda L.) and Scots pine (Pinus sylvestris L.)

The effects of enhanced UV‐B radiation on the needle anatomy of loblolly pine (Pinus taeda L.) and Scots pine (Pinus sylvestris L.) were studied in the field under supplemental UV‐B radiation supplied by a modulated irradiation system. The supplemental UV‐B levels were designed to simulate either a 16 or 25% loss of stratospheric ozone over College Park, Maryland. Enhanced UV‐B radiation caused different responses in these two species. The needles of loblolly pine had larger amounts of tannin in the lumen of epidermal cells and more wall‐bound phenolics in the outer epidermal walls of UV‐B‐treated needles, whereas the most pronounced effect on Scots pine needles was increased cutinization. In both species, the outer epidermal cell walls thickened and the needle cross‐sectional and mesophyll areas decreased (statistically significantly only in Scots pine). This suggests that more carbon may have been allocated to the protection mechanisms at the expense of photosynthetic area. The difference in response between these species suggests that the response to UV‐B radiation is not mediated by a single mechanism and that no generalization with regard to the effects of UV‐B on conifers can be made.     

棉花萌发期和苗期耐盐性评价及耐盐指标筛选

于2010年在江苏南京农业大学人工气候室内进行砂培试验,采用不同浓度的NaCl水溶液模拟盐胁迫,以多项指标盐害系数隶属函数值和总隶属函数值为依据,比较了13个棉花品种萌发期和苗期的耐盐性,并进行聚类分析.结果表明:150mmol·L-1NaCl是进行棉花耐盐性鉴定的适宜盐浓度.棉花的耐盐性在生育期和品种间表现不同:中棉所44和中棉所177是在萌发期和苗期均表现稳定的耐盐品种,表现稳定但不耐盐品种有中棉所102、苏棉12号和泗棉3号,表现稳定且中等耐盐品种有中棉所103、德夏棉1号和美棉33B;发芽率、发芽势、发芽指数、活力指数和鲜质量的盐害系数可以作为棉花萌发期耐盐鉴定指标,株高、叶片伸展速率、地上部干质量、根系干质量、根系活力和净光合速率的盐害系数可以作为棉花苗期耐盐鉴定指标.     

两个棉花Rac蛋白基因的克隆与表达分析

为研究棉花纤维起始和伸长的分子机理,在棉花纤维EST序列分析的基础上,从棉花纤维中扩增并克隆了2个棉花Rac蛋白的cDNA基因,分别命名为GhRacA和GhRacB。GhRacA cDNA长959bp,推测的编码蛋白包含211个氨基酸。GhRacB cDNA长920bp,编码195个氨基酸的蛋白。GhRacA和GhRacB蛋白均含有GTP／GDP结合和激活区域、Effector区和碱性氨基酸区。GhRacB的C末端有保守的异戊烯基化位点CSIL,而GhRacA没有明显的异戊烯基化位点。序列比较分析表明,GhRacA和GhRacB是2个新的棉花Rac蛋白。RT-PCR分析表明,GhRacA和GhRacB在根、下胚轴、茎、叶和纤维中都有表达,但均在棉花纤维起始和伸长时期有优势表达,推测2个基因在棉花纤维的早期发育中可能有重要的功能。