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Dedifferentiating initiation and embryogenesis from freshly-isolated microspores of barley
Xiangrong Guo,Jiankang Jing,Han Hu
Science China Life Sciences , 1997, DOI: 10.1007/BF02879095
Abstract: By using DNA-specific fluorescent dye and a confocal laser scanning microscope, the present study was designed to investigate the cytological characteristics of dedifferentiating initiation during pretreatment and embryogenesis during culture in freshly-isolated microspores of barley, and the difference in main developmental pathway between freshly-isolated and cold-treated microspores. The results revealed that (i) freshly-isolated microspores started the initiation within 12 h of mannitol pretreatment, whose main cytological characteristics were that: cell volume was obviously extended; the volume of nuclei and nucleoli were also greatly increased; nucleoli were extremely clear and highly condensed; N/C ratio was very high; (ii) all the pretreatment methods led to the initiation of the microspores, thus triggering the embryogenic process; (iii) pretreatment methods influenced the main developmental pathway of microspores by changing the pattern of the first mitosis. The cold-treated microspores formed main developmental pathway via A patterns, but freshly-isolated microspores via B pattern.
Dedifferentiating initiation and embryogenesis from freshly-isolated microspores of barley
Guo Xiangrong,Jing Jiankang,Hu Han,
郭向荣
,景建康,胡含

中国科学C辑(英文版) , 1997,
Abstract: By using DNA-specific fluorescent dye and a confocal laser scanning microscope, the present study was designed to investigate the cytological characteristics of dedifferentiating initiation during pretreatment and em-bryogenesis during culture in freshly-isolated microspores of barley, and the difference in main developmental pathway between freshly-isolated and cold-treated microspores. The results revealed that ( i ) freshly-isolated microspores started the initiation within 12 h of mannitol pretreatment, whose main cytological characteristics were that: cell vol-ume was obviously extended; the volume of nuclei and nucleoli were also greatly increased; nucleoli were extremely clear and highly condensed; N/C ratio was very high; ( ii ) all the pretreatment methods led to the initiation of the mi-crospores, thus triggering the embryogenic process; ( iii ) pretreatment methods influenced the main developmental pathway of microspores by changing the pattern of the first mitosis. The cold-treated microspores formed main devel-opmental pathway via A patterns, but freshly-isolated microspores via B pattern.
In Vitro Silencing of Brugia malayi Trehalose-6-Phosphate Phosphatase Impairs Embryogenesis and In Vivo Development of Infective Larvae in Jirds  [PDF]
Susheela Kushwaha,Prashant Kumar Singh,Mohd. Shahab,Manisha Pathak,Shailja Misra Bhattacharya
PLOS Neglected Tropical Diseases , 2012, DOI: 10.1371/journal.pntd.0001770
Abstract: Background The trehalose metabolic enzymes have been considered as potential targets for drug or vaccine in several organisms such as Mycobacterium, plant nematodes, insects and fungi due to crucial role of sugar trehalose in embryogenesis, glucose uptake and protection from stress. Trehalose-6-phosphate phosphatase (TPP) is one of the enzymes of trehalose biosynthesis that has not been reported in mammals. Silencing of tpp gene in Caenorhabditis elegans revealed an indispensable functional role of TPP in nematodes. Methodology and Principal Findings In the present study, functional role of B. malayi tpp gene was investigated by siRNA mediated silencing which further validated this enzyme to be a putative antifilarial drug target. The silencing of tpp gene in adult female B. malayi brought about severe phenotypic deformities in the intrauterine stages such as distortion and embryonic development arrest. The motility of the parasites was significantly reduced and the microfilarial production as well as their in vitro release from the female worms was also drastically abridged. A majority of the microfilariae released in to the culture medium were found dead. B. malayi infective larvae which underwent tpp gene silencing showed 84.9% reduced adult worm establishment after inoculation into the peritoneal cavity of na?ve jirds. Conclusions/Significance The present findings suggest that B. malayi TPP plays an important role in the female worm embryogenesis, infectivity of the larvae and parasite viability. TPP enzyme of B. malayi therefore has the potential to be exploited as an antifilarial drug target.
影响甘蓝小孢子DH植株生长的几个因素
Influence of Several Factors on Growth of DH Plants Regenerated from Microspores in Cabbage
 [PDF]

,,张恩慧,程永安,许忠民,王改改
- , 2016, DOI: 10.7606/j.issn.1004-1389.2016.04.017
Abstract: 旨在筛选甘蓝小孢子DH植株健壮生长的最适培养基和光照强度。为缩短小孢子DH植株培养时间并为获得健壮植株创造条件,从而加快甘蓝DH系育种进程。以甘蓝‘秦甘50’为材料进行游离小孢子培养至诱导分化出不定芽,转接到NAA质量浓度不同的2种培养基上(1/2MS+0、0.1、0.2、0.3 mg/L NAA和MS+0.1 mg/L NAA),不同光照强度[40、60、80、100 μmol/(m ·s)]及添加不同质量浓度NH4NO3(0、0.4、0.8、1.2 g/L)培养,研究DH植株的生长势,获得甘蓝小孢子DH植株最适培养基和培养条件。结果表明,在NAA质量浓度不同的培养基中,1/2MS +0.1 mg/L NAA培养基中小孢子DH植株生长速度最显著,生根快、植株长势强;在80 μmol/(m ·s)光照强度下培养小孢子DH植株净生长高度显著,10.3 d再生出根系,根质量较好,叶片大而厚;MS培养基中添加质量浓度为0.4 g/L的NH4NO3,小孢子DH植株生长健壮,35 d内植株净生长高度最大为56.25 mm。以上结果说明,甘蓝小孢子DH植株快速健壮生长的适宜培养基为1/2MS+0.1 mg/L NAA+8 g/L琼脂+30 g/L蔗糖;在80 μmol/(m ·s)光照强度下或MS培养基中添加质量浓度为0.4 g/L的NH4NO3均能够促进甘蓝小孢子DH植株根茎叶生长旺盛。
Aims to screen the most appropriate medium and illumination for DH plants regenerated from microspore in cabbage and to shorten culture time of obtaining robust plants, which will accelerate the breeding process of cabbage double haploid. Microspores of ‘Qin-gan 50’ cabbage was cultured until emergence of adventitious buds which then transferred to two kinds of mediums (1/2MS+0,0.1,0.2,0.3 mg/L NAA and MS+0.1 mg/L NAA), adding different amount of NH4NO3(0,0.4,0.8,1.2 g/L) in MS medium and cultured at different illumination [40,60,80,100 μmol/(m ·s)] . Then the growth vigor of the DH plants were studied and the most appropriate medium and culture conditions could be gotten. The DH plants grown in 1/2MS +0.1 mg/L NAA medium had the fastest growth rate and strong growth vigor, rooting fast;cultured in the illumination of 80 μmol/(m ·s),the DH plants had the most remarkable growth speed, better roots and leaves, which rooted in 10.3 days;the DH plants in MS medium with added 0.4 g/L NH4NO3 had the maximum value 56.25 mm of net growth height in 35 days. In conclusion, the appropriate medium for the DH plants regenerated from microspore in cabbage is 1/2MS+0.1 mg/L NAA+8 g/L Agar+30 g/L Sugar;cultured in the illumination of 80 μmol/(m ·s) or transferred into the MS medium with added 0.4 g/L NH4NO3, the DH plants grown well with robust roots, stems and leaves.
Efficient Somatic Embryogenesis and Organogenesis of Self-Pollination Artemisia annua Progeny and Artemisinin Formation in Regenerated Plants  [PDF]
Fatima Alejos-Gonzalez, Kelly Perkins, Malcolm Isaiah Winston, De-Yu Xie
American Journal of Plant Sciences (AJPS) , 2013, DOI: 10.4236/ajps.2013.411274
Abstract: To enhance the understanding of artemisinin biosynthesis, we have successfully bred self-pollination Artemisia annua plants. Here, we report efficient somatic embryogenesis and organogenesis of self-pollination plants and artemisinin formation in regenerated plants. The first through sixth nodal leaves of seedlings are used as explants. On agar-solidified MS basal medium supplemented with TDZ (0.6 mg/l) and IBA (0.1 mg/l), all explants after inoculation of less than 3 weeks start to form embryogenic calli, which further produce globular, torpedo, heart and early cotyledon embryos. In all six positional leaves, explants from the sixth leaf show the rapidest responses to induction of embryogenic calli and somatic embryos. On this medium, somatic embryos continuously develop into adventitious buds, which can form adventitious roots on a rooting medium containing NAA (0.5 mg/l). Meanwhile, on agar-solidified MS basal medium supplemented with BAP (1 mg/l) and NAA (0.05 mg/l), approximately 100% of explants from leaves #3-6 form calli in less than 3 weeks of inoculation and adventitious buds via organogenesis in 3-4 weeks. In all six positional leaves, explants from the sixth leaf exhibit the rapidest response to induction of calli and adventitious buds. Nearly 100% adventitious buds can form adventitious roots on the rooting medium. Regenerated plants from both somatic embryogenesis and organogenesis complete self-pollination to produce seeds in 80-90 days of growth in growth chamber. LC-ESI-MS analysis demonstrates that regenerated plants biosynthesize artemisinin. These results show the highly efficient regeneration capacity of self-pollination
Isolation and culture of soybean (Glycine max L. Merrill) microspores and pollen grains
Rodrigues, Lia Rosane;Forte, Bianca de Camargo;Bodanese-Zanettini, Maria Helena;
Brazilian Archives of Biology and Technology , 2006, DOI: 10.1590/S1516-89132006000500002
Abstract: in the last three decades, research on soybean microspore embryogenesis was restricted to anther culture, which presents limitations such as the small number of responsive microspores and the high embryogenic potential of sporophytic tissues. therefore, a sequence of studies was performed to establish appropriate conditions for the isolation and culture of soybean microspores and pollen grains as an alternative to anther culture. first, a pollen and microspore isolation technique was developed using floral buds from four soybean cultivars (bragg, ias 5, mg/br-46 conquista and brsmt uirapuru). this technique allowed the establishment of cultures with satisfactory density and characteristics. subsequently, different culture conditions were tested. although b5 and ms media have been currently recommended for soybean anther culture, the best result was obtained in pta-15 modified medium, with the formation of enlarged microspores and 0.4% of multicellular pollen grains in the cultivar brsmt uirapuru.
Polyamines and Carcinogenesis
Uriel Bachrach
Acta Facultatis Medicae Naissensis , 2012,
Abstract: The naturally occurring polyamines, spermine, spermidine and the diamine putrescine are widespread in nature. They have been implicated in growth and differentiation processes. In 1967, we reported that cancer cells are rich in polyamines. Subsequently, it has been shown that polyamines are released from cancer cells and may be detected in body fluids such as urine, blood and cerebrospinal fluids. It has also been demonstrated that the increase in cellular polyamine levels is an early and an obligatory event in the process of malignant transformation. This increase in cellular polyamine concentration is due to the activation of ornithine decarboxylase (ODC), which catalyses the rate limiting step in polyamine synthesis by converting ornithine to putrescine. Assays of urinary and blood polyamines have been used to detect cancer and to determine the success of therapy. A sensitive, rapid, chemiluminescence-based method for the determination of diamines and polyamines was developed and 2.000 urine samples were tested. An interesting "gene therapy" system for injecting amine oxidases into normal and transformed cells was developed as follows: serum amine oxidase and porcine kidney damine oxidase were trapped within reconstituted Sendai virus envelopes. Chick or rat fibroblasts, transformed by Rous sarcoma virus, were more susceptible to the injected enzymes, compared to the normal culture, when macromolecular synthesis was tested. An in vitro chemosensitivity assay for the testing of the sensitivity of cancer cells from individual patients ("tailored treatment") was also developed. All these studies stress the importance of polyamines in carcinogenesis.
Induction of symmetrical nucleus division and multi-nucleate structures in microspores of eggplant (Solanum melongena L.) cultured in vitro
Bal, Ugur;Ellialtioglu, Sebnem;Abak, Kazim;
Scientia Agricola , 2009, DOI: 10.1590/S0103-90162009000400016
Abstract: a modification of a protocol used to induce tobacco microspore embryogenesis was tested in eggplant (solanum melongena l.). in tobacco, uninucleate microspores are subjected to stress treatment by culturing in mannitol containing "b" medium at 33oc for six days. the microspores are then transferred to maltose containing at3 medium for further development. in the experiment presented here late uninucleate and bi-nucleate microspores of the eggplant cultivar bambino were pre-cultured in b medium and then incubated at +4oc, 25oc and 33oc, respectively, for two days. after the pre-treatments, microspore cultures were transferred to at3 medium containing 0.25 m maltose and maintained at 25oc in the dark. presence of symmetrical division and multinucleate structures was checked with dapi staining of the nucleus after one and two weeks. symmetrical division of the nucleus and multinucleate structures were observed only in uni-nucleate microspores pre-treated at 33oc for two days. the frequency of multinucleate structures was 19.4% under these conditions. we demonstrated that eggplant is responsive to the modified tobacco protocol in the production of symmetrically division and multinucleate structures. these results may be used as a basis for adaptation fully of the tobacco system in eggplant.
Embryogenesis of Isolated-microspore in Cabbage (Brassica oleracea var.capitata)
结球甘蓝游离小孢子胚胎发生

FANG Shu-Gui,ZENG Xiao-Ling,ZHU Chao-Hui,LIN Bi-Ying,CHEN Wen-Hui,LIAO Xiao-Zhen,
方淑桂
,曾小玲,朱朝辉,林碧英,陈文辉,廖晓珍

植物科学学报 , 2005,
Abstract: Factors affecting microspore embryogenesis in cabbage was intensively investigated(using) cultivar,'Qiangxia'.Results showed that the optimum period for donor flower buds collection was before full bloom of the plants.Embryoids could develop from microspores of uninucleate stage to binucleate stage.Medium with 17% sucrose at the first days of culture was favorable for the vitality of microspores,but medium with 14% sucrose was favorable for the embryogenesis after 3 days of culture.It was found that the embryogenesis ability was greatly improved by adding low sucrose concentration(11%) medium into the medium with high sucrose concentration(17%) after 3 days of culture.In this case,the embryogenesis frequency increased 282.4% and 126.1% respectively compared with microspores cultured consistently in medium with 14% sucrose or changing the medium completely.
Nuclear Fusion during Early Stage of Microspore Embryogenesis Indicates Chromosome Doubling in Wheat (Triticum aestivum)  [PDF]
Roland Griggs, Ming Y. Zheng
American Journal of Plant Sciences (AJPS) , 2016, DOI: 10.4236/ajps.2016.73043
Abstract: Studies of barley and maize indicate that chromosome doubling occurs via nuclear fusion during an early stage of microspore embryogenesis, but the time and mechanism by which chromosome doubling occurs in bread wheat (Triticum aestivum) remains undetermined. The purpose of this study was to determine the relative time during induction culture when chromosome doubling may occur in wheat, and to identify early indicators for doubled haploid microspores. Microspore nuclei were stained with 4’,6-diamidino-2-phenylindole (DAPI) and observed under a fluorescent microscope on the day of isolation, three days after isolation, and six days after isolation. The change in the percentage of microspores containing a single small nucleus, two small nuclei, a single enlarged nucleus, and three or more nuclei was then tracked throughout the six-day period. Ploidy levels were estimated by determining the cross-sectional area and number of nucleoli in microspores containing small and large nuclei then comparing the results of each respective cell-type. The percentage of microspores containing enlarged nuclei increased throughout the six-day test period, and the percentage of binucleated microspores containing small nuclei decreased. Comparison of the changes in average percentage of microspores containing a single small nucleus, binucleated microspores, microspores containing a single large nucleus, and multinucleate microspores on days 0, 3, and 6 indicates that nuclei classified as “small” are likely haploids and nuclei classified as “large” are doubled haploids. The percentage of microspores with enlarged nucleus (nuclei) during the first six days of induction culture could be used as an early indicator for the frequency of chromosome doubling in wheat microspore culture.
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