Analysis of the barley bract suppression gene Trd1

A typical barley (Hordeum vulgare) floret consists of reproductive organs three stamens and a pistil, and non-reproductive organs-lodicules and two floral bracts, abaxial called 'lemma' and adaxial 'palea'. The floret is subtended by two additional bracts called outer or empty glumes. Together these organs form the basic structural unit of the grass inflorescence, a spikelet. There are commonly three spikelets at each rachis (floral stem of the barley spike) node, one central and two lateral spikelets. Rare naturally occurring or induced phenotypic variants that contain a third bract subtending the central spikelets have been described in barley. The gene responsible for this phenotype was called the THIRD OUTER GLUME1 (Trd1). The Trd1 mutants fail to suppress bract growth and as a result produce leaf-like structures that subtend each rachis node in the basal portion of the spike. Also, floral development at the collar is not always suppressed. In rice and maize, recessive mutations in NECK LEAF1 (Nl1) and TASSEL SHEATH1 (Tsh1) genes, respectively, have been shown to be responsible for orthologous phenotypes. Fine mapping of the trd1 phenotype in an F-3 recombinant population enabled us to position on the long arm of chromosome 1H to a 10 cM region. We anchored this to a conserved syntenic region on rice chromosome Os05 and selected a set of candidate genes for validation by resequencing PCR amplicons from a series of independent mutant alleles. This analysis revealed that a GATA transcription factor, recently proposed to be Trd1, contained mutations in 10 out of 14 independent trd1 mutant alleles that would generate non-functional TRD1 proteins. Together with genetic linkage data, we confirm the identity of Trd1 as the GATA transcription factor ortholog of rice Nl1 and maize Tsh1 genes.

NAA and STS effects on bract survival time, carbohydrate content, respiration rate and carbohydrate balance of potted Bougainvillea spectabilis Willd

The aims of this work were to deepen the knowledge on the physiology of bract abscission in Bougainvillea spectabilis ‘Killie Campbell’ plants, in what relates to respiration and carbon balance. Using the effects induced by Silver Thiosulphate (STS) and/or Naphtalene Acetic Acid (NAA, at high concentration: 500 mg.l-1) on bract abscission under interior conditions, the relationship between bract survival time (longevity) and, respiration rate or carbohydrate levels, was investigated. Treatments that included NAA were the ones that reduced significantly bract abscission. Unexpectedly, the higher the levels of bract soluble and total carbohydrates, measured at day 10 postproduction (PP), the higher the abscission of bracts. These results show, for the first time, that abscission can positively correlate with non structural carbohydrates levels in the organ that abscise. Bract respiration rate was significantly affected by treatment and postproduction day (PP). Treatments that had higher bract respiration rates (WATER and STS) also had higher levels of non structural carbohydrates in the bracts. Bract respiration rate decreased from day 10 to day 17 PP by approximately 50% (on average of all treatments) and was negatively correlated with bract survival time. In the carbon balance per gram of bract dry weight, the treatments WATER and STS, showed the largest decrease in the content of total carbohydrates and had the highest consumption of carbohydrates through respiration. So, these were the bracts that needed to import a higher amount of carbohydrates per gram of bract dry weight. In the carbon balance for the whole mass of bracts and adjacent stems in an average plant, the treatments WATER and STS continued to allow for the largest decreases in total carbohydrate during postproduction. However, and contradicting the results per gram of bract dry weight, the highest total consumption of carbohydrates by respiration was obtained for the NAA and STS+NAA treatments. It makes sense that bracts that last longer have lower individual carbon consumption while, at the plant level, the increased number of remaining bracts causes a higher overall expenditure. Respiration rate has been used as an indicator of flower longevity, this correlation is here extended for the flower+bract system. Plants that had higher bract respiration rates, most probably, had a higher flow of carbohydrates through the bracts (and flowers), which, in the end, was sensed as a higher carbohydrate level.

The transformation of banana with potential virus resistance genes

One approach to reducing the yield losses caused by banana viral diseases is the use of genetic engineering and pathogen-derived resistance strategies to generate resistant cultivars. The development of transgenic virus resistance requires an efficient banana transformation method, particularly for commercially important 'Cavendish' type cultivars such as 'Grand Nain'. Prior to this study, only two examples of the stable transformation of banana had been reported, both of which demonstrated the principle of transformation but did not characterise transgenic plants in terms of the efficiency at which individual transgenic lines were generated, relative activities of promoters in stably transformed plants, and the stability of transgene expression. The aim of this study was to develop more efficient transformation methods for banana, assess the activity of some commonly used and also novel promoters in stably transformed plants, and transform banana with genes that could potentially confer resistance to banana bunchy top nanovirus (BBTV) and banana bract mosaic potyvirus (BBrMV). A regeneration system using immature male flowers as the explant was established. The frequency of somatic embryogenesis in male flower explants was influenced by the season in which the inflorescences were harvested. Further, the media requirements of various banana cultivars in respect to the 2,4-D concentration in the initiation media also differed. Following the optimisation of these and other parameters, embryogenic cell suspensions of several banana (Musa spp.) cultivars including 'Grand Nain' (AAA), 'Williams' (AAA), 'SH-3362' (AA), 'Goldfinger' (AAAB) and 'Bluggoe' (ABB) were successfully generated. Highly efficient transformation methods were developed for both 'Bluggoe' and 'Grand Nain'; this is the first report of microprojectile bombardment transformation of the commercially important 'Grand Nain' cultivar. Following bombardment of embryogenic suspension cells, regeneration was monitored from single transfom1ed cells to whole plants using a reporter gene encoding the green fluorescent protein (gfp). Selection with kanamycin enabled the regeneration of a greater number of plants than with geneticin, while still preventing the regeneration of non-transformed plants. Southern hybridisation confirmed the neomycin phosphotransferase gene (npt II) was stably integrated into the banana genome and that multiple transgenic lines were derived from single bombardments. The activity, stability and tissue specificity of the cauliflower mosaic virus 358 (CaMV 35S) and maize polyubiquitin-1 (Ubi-1) promoters were examined. In stably transformed banana, the Ubi-1 promoter provided approximately six-fold higher p-glucuronidase (GUS) activity than the CaMV 35S promoter, and both promoters remained active in glasshouse grown plants for the six months they were observed. The intergenic regions ofBBTV DNA-I to -6 were isolated and fused to either the uidA (GUS) or gfjJ reporter genes to assess their promoter activities. BBTV promoter activity was detected in banana embryogenic cells using the gfp reporter gene. Promoters derived from BBTV DNA-4 and -5 generated the highest levels of transient activity, which were greater than that generated by the maize Ubi-1 promoter. In transgenic banana plants, the activity of the BBTV DNA-6 promoter (BT6.1) was restricted to the phloem of leaves and roots, stomata and root meristems. The activity of the BT6.1 promoter was enhanced by the inclusion of intron-containing fragments derived from the maize Ubi-1, rice Act-1, and sugarcane rbcS 5' untranslated regions in GUS reporter gene constructs. In transient assays in banana, the rice Act-1 and maize Ubi-1 introns provided the most significant enhancement, increasing expression levels 300-fold and 100-fold, respectively. The sugarcane rbcS intron increased expression about 10-fold. In stably transformed banana plants, the maize Ubi-1 intron enhanced BT6.1 promoter activity to levels similar to that of the CaMV 35S promoter, but did not appear to alter the tissue specificity of the promoter. Both 'Grand Nain' and 'Bluggoe' were transformed with constructs that could potentially confer resistance to BBTV and BBrMV, including constructs containing BBTV DNA-1 major and internal genes, BBTV DNA-5 gene, and the BBrMV coat protein-coding region all under the control of the Ubi-1 promoter, while the BT6 promoter was used to drive the npt II selectable marker gene. At least 30 transgenic lines containing each construct were identified and replicates of each line are currently being generated by micropropagation in preparation for virus challenge.

Detection of banana bunchy top virus in virus-infected plants using polymerase chain reaction

Polymerase chain reaction (PCR) was developed for the detection of Banana bunchy top virus (BBTV) at maximum after 210 min and at minimum after 90 min using Pc-1 and Pc-2, respectively. PCR detection of BBTV in crude sap indicated that the freezing of banana tissue in liquid nitrogen (LN2) before extraction was more effective than using sand as the extraction technique. BBTV was also detected using PCR assay in 69 healthy and diseased plants using Na-PO4 buffer containing 1 % SDS. PCR detection of BBTV in nucleic acid extracts using seven different extraction buffers to adapt the use of PCR in routine detection in the field was studied. Results proved that BBTV was detected with high sensitivity in nucleic acid extracts more than in infectious sap. The results also suggested the common aetiology for the BBTV by the PCR reactions of BBTV in nucleic acid extracts from Australia, Burundi, Egypt, France, Gabon, Philippines and Taiwan. Results also proved a positive relation between the Egyptian-BBTV isolate and abaca bunchy top isolate from the Philippines, but there no relation was found with the Cucumber mosaic cucumovirus (CMV) isolates from Egypt and Philippines and Banana bract mosaic virus (BBMV) were found.

Abacá mosaic virus: a distinct strain of Sugarcane mosaic virus

Abacá mosaic virus (AbaMV) is related to members of the sugarcane mosaic virus subgroup of the genus Potyvirus. The ~2 kb 3′ terminal region of the viral genome was sequenced and, in all areas analysed, found to be most similar to Sugarcane mosaic virus (SCMV) and distinct from Johnsongrass mosaic virus (JGMV), Maize dwarf mosaic virus (MDMV) and Sorghum mosaic virus (SrMV). Cladograms of the 3′ terminal region of the NIb protein, the coat protein core and the 3′ untranslated region showed that AbaMV clustered with SCMV, which was a distinct clade and separate from JGMV, MDMV and SrMV. The N-terminal region of the AbaMV coat protein had a unique amino acid repeat motif different from those previously published for other strains of SCMV. The first experimental transmission of AbaMV from abacá (Musa textilis) to banana (Musa sp.), using the aphid vectors Rhopalosiphum maidis and Aphis gossypii, is reported. Polyclonal antisera for the detection of AbaMV in western blot assays and ELISA were prepared from recombinant coat protein expressed in E. coli. A reverse transcriptase PCR diagnostic assay, with microtitre plate colourimetric detection, was developed to discriminate between AbaMV and Banana bract mosaic virus, another Musa-infecting potyvirus. Sequence data, host reactions and serological relationships indicate that AbaMV should be considered a distinct strain of SCMV, and the strain designation SCMV-Ab is suggested.

Evaluación del crecimiento y producción de turiones de espárrago verde (Asparagus officinalis L.) producidos bajo dos sistemas de inicio, arañas y plantines, en Azul, Buenos Aires

En dos plantaciones de espárrago verde (Asparagus officinalis L.) en el centro de la Pcia. de Buenos Aires, del cultivar UC 157 F2, iniciadas con arañas y plantines, durante su primer año de cosecha, se determinó la tasa de elongación diaria y total hasta el momento de la ramificación de los turiones, el número de días hasta alcanzar el largo exportable (22.5 cm) y los días hasta el momento de la ramificación con el objetivo de estudiar su comportamiento en la zona centro de la provincia de Buenos Aires. Las mediciones se efectuaron durante seis semanas, desde el 26 de septiembre hasta el 4 de noviembre de 2005, desde la emergencia del turión hasta el día de su ramificación (rama de 5 mm sobre su bráctea). La elongación máxima hasta el día de ramificación en promedio y para ambos sistemas de inicio, varió 2 desde 35.3 cm al principio de la cosecha hasta 22.8 cm al final de la misma. Las tasas de elongación relativas variaron en promedio para ambos sistemas de inicio, entre 0.11 cm cm-1 d-1 y 0.38 cm cm-1 d-1 dependiendo de la temperatura del aire. Al mismo tiempo las parcelas constituyentes del ensayo fueron sometidas a una cosecha comercial durante las seis semanas de evaluación. Los espárragos cosechados constituyeron el rendimiento total (kg ha-1) para cada sistema de inicio entre los cuales no se encontraron diferencias estadísticamente significativas. Este estudio permitiría explicar las diferencias de rendimiento comercial para un mismo cultivar en diferentes zonas de producción. Se concluye que para una zona dada, la elongación y sobre todo la ramificación, son variables prioritarias para la determinación del potencial productivo de un cultivar, debiendo tener en cuenta las condiciones de temperatura en las cual se pretende producir este cultivo y el mercado de destino de la producción debido a las mercadas preferencias entre sus consumidores.

柏科植物雌球果发育的研究

在裸子植物系统学研究中，雌球果的形态与结构是最重要的性状之一。山于不同学者对裸子植物雌性生殖构造的形态学本质有着不同的见解，从而导致对该类群植物系统发育研究结果的差异。柏科在裸子植物中是唯一一个广布于南北两半球的科，前人对该科雌球果发育的研究涉及很少，为了进一步了解柏科植物雌球果的形态学本质，并为裸子植物传粉生物学和系统学的研究提供重要资料，本文选取了分布于北半球柏科中的7个属14个种，应用扫描电镜、半薄切片、石蜡切片及整体透明等方法，对其雌球果各部分的形态发生与发育过程作了详尽的研究，同时对传粉前后雌球果中的维管系统作了深入的分析。主要结果归纳如下： 1)雌球花苞片的形态、结构和排列方式等与叶相似;但传粉后，随着苞片基部的居间生长，其形态结构变得多样化： 2)在柏科了个属中，均未观察到种鳞发育，成熟球果的苞鳞得到充分发育; 3)各苞片腋内的胚珠发育过程，几乎是同时发育或呈向顶发育过程; 4)在圆柏属和柏木属球果顶端的苞片腋内也有胚珠发育，其它各属雌球花顶端的1-2对苞片腋内均没有胚珠发育; 5)在柏木属中，每一苞片腋内形成的胚珠数目较多，一般排成2-3行，同～苞片腋内的胚珠呈向基发育，同一行胚珠为离心发育; 6)圆柏属的雌球果发育呈现多种式样，其中包括由数枚侧生胚珠过渡到一枚顶生胚珠等情况; 7)柏科植物雌球花的维管系统在传粉前较为简单，即在苞片内仅具单一的维管束延伸到顶端，与营养芽相似。传粉后随着苞片基部的居间生长，其维管系统也随之复杂化。主要是在苞鳞内有新的维管束形成，一般是在苞鳞中间维管束的两侧各发出一束维管束，这两束维管束由球果轴发出的位置略高于中间一束，并在苞鳞基部迅速分枝，形成不同的排列式样。 成熟球果维管系统的排列方式与球果和苞鳞的形态有关，通常可分为三类：(1)具球形球果的种类，包括圆柏属、扁柏属、柏木属和福建柏属，其苞鳞内辐射状分枝的维管束形成两组，即～个具有木质部倒转的近轴系列和一个维管束J下常排列的远轴系列;(2)在苞鳞中只形成一组倒转的维管束，如侧柏属和翠柏属：(3)崖柏属雌球果的苞鳞中没有倒转的维管束形成。 观察结果表明：柏科植物雌球果均为复轴性构造，其球果中胚珠着生在球果轴上是由于苞片腋内种鳞退化的结果，因而不支持柏科植物雌球果的种鳞与苞鳞完全合生的观点。 通过对柏科植物雌球果发育的研究结果，也为探讨松杉类植物雌球果的演化趋势提供了重要线索。长期以来，众多研究者均认为，松杉类植物雌球果的演化趋势是种鳞和苞鳞逐渐合并的过程，但从柏科植物雌球果苞鳞腋内种鳞退化现象的发现表明，松杉类植物雌球果的演化趋势可能是一种种鳞逐渐退化的过程。柏科的种鳞已完全退化，而苞鳞则相对发育。 依据雌球果发育中胚珠发育顺序及苞鳞数目等特征，认为福建柏属最原始，而圆柏属则较为进化，其它各属则介于上述两属之间。 江泽平和王豁然(1997)提出，柏木亚科和澳洲柏亚科的主要区别在于球果可育苞鳞的着生位置，如前者球果顶部苞鳞没有胚珠发育，而后者则是球果顶部苞鳞腋内有胚珠发育。这一观点显然与柏木属和圆柏属的多数雌球果顶部的苞鳞腋内有种子产生的情况不相符合。 依据本文的研究的结果及前人对柏科和相近类群的研究资料，赞同保留独立的柏科和杉科;另外，以往侧柏曾被置于崖柏属中，但依据其雌球果苞鳞中有倒转的维管束发育，崖柏属的崖柏和北美香柏雌球果的苞鳞中均无倒转的维管束发育，我们支持其独立为侧柏属。

Inflorescence and floral development of Dahlstedtia species (Leguminosae: Papilionoideae: Millettieae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morfoanatomia das brácteas em Heliconia (Heliconiaceae) ocorrentes no Estado de São Paulo, Brasil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Homologies of the flower and inflorescence in the early-divergent grass anomochloa (poaceae)

Premise of the study: The grass subfamily Anomochlooideae is phylogenetically significant as the sister group to all other grasses. Thus, comparison of their structure with that of other grasses could provide clues to the evolutionary origin of these characters. Methods: We describe the structure, embryology, and development of the flower and partial inflorescence of the monotypic Brazilian grass Anomochloa marantoidea. We compare these features with those of other early-divergent grasses such as Pharus and Streptochaeta and closely related Poales such as Ecdeiocolea. Key results: Anomochloa possesses several features that are characteristic of Poaceae, notably a scutellum, a solid style, reduced stamen number, and an ovary with a single ovule that develops into a single indehiscent fruit. Interpretation of floral patterning in Anomochloa is problematic because the ramification pattern of the florets places the bracts and axes in unusual positions relative to the primary inflorescence axis. Our study indicates that there is a single abaxial carpel in Anomochloa, probably due to a cryptic type of pseudomonomery in Anomochloa that resembles the pseudomonomery of other grasses. On the other hand, the Anomochloa flower differs from the typical grass flower in lacking lodicules and possessing four stamens, in contrast with the tristaminate condition that characterizes many other grasses. Conclusions: Using the median part of the innermost bract as a locator, we tentatively homologize the inner bract of the Anomochloa partial inflorescence with the palea of other grasses. In this interpretation, the pattern of monosymmetry due to stamen suppression differs from that of Ecdeiocolea. © 2012 Botanical Society of America.

Caracterização da variabilidade morfoagronômica de maracujazeiro (Passiflora cincinnata Mast.) no semi-árido brasileiro

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)