Green Fluorescent Protein (GFP) in Vector Systems Played Sense Role of Epigenetic in Plants

The green fluorescent protein (GFP) of jellyfish (_Aequorea victoria_) has significant advantages over other reporter genes, because expression can be detected in living cells without any substrates. Recently, epigenetic phenomena are important to consider in plant biotechnology experiments for elucidate unknown mechanism. Therefore, soybean immature cotyledons were generated embryogenesis cells and engineered with two different gene constructs (pHV and pHVS) using gene gun method. Both constructs contain a gene conferring resistance to hygromycin (_hpt_) as a selective marker and a modified glycinin (11S globulin) gene (_V3-1_) as a target. However, sGFP(_S65T_) as a reporter gene was used only in pHVS as a reporter gene for study the relation between using sGFP(_S65T_) and gene silencing phenomena. Fluorescence microscopic was used for screening after the selection of hygromycin, identified clearly the expression of sGFP(_S65T_) in the transformed soybean embryos bombarded with the pHVS construct. Protein analysis was used to detect gene expression overall seeds using SDS-PAGE. Percentage of gene down regulation was highly in pHV construct compared with pHVS. Thus, sGFP(_S65T_) as a reporter gene in vector system may be play useful role for transgenic evaluation and avoid gene silencing in plants for the benefit of plant transformation system

Effect of Growth Regulators on In Vitro Morphogenic Response of Boscia senegalensis (Pers.) Lam. Poir. Using Mature Zygotic Embryos Explants

The percent study describes the in vitro responses of mature zygotic embryos of Boscia senegalensis to different concentrations (0.0–5.0 mg/L) of 6-benzyladnine (BA), Thidiazuron (TDZ), α-Naphthalene acetic acid (NAA), and 2, 4-Dichlorophenoxyacetic acid (2, 4-D) supplemented on Murashige and Skoog medium (MS). The plant growth regulators (PGRs) were considerably affected the morphogenetic responses. BA produced adventitious shoots through two ways: direct organogenesis and auxiliary shoot formation. Both 2, 4-D and TDZ tend to produce callus, whereas NAA improve the development of embryos to seedlings. Maximum number of shoots/explant (14.8 ± 0.6) was obtained on MS medium supplemented with 3.0 mg/L BA. 67.0% of excised shoots were rooted either on 1/2 MS medium augmented with or without 0.25 mg/L IBA. The highest number of roots (1.2 ± 0.4) and root length (0.5 ± 0.2 cm) was produced on 0.25 mg/L IBA-containing medium. Regenerated plants were successfully acclimatized and transferred to the green house with 70% survival rate. All the plants appeared morphologically uniform with normal growth pattern. A rapid (30 days), efficient and without subculturing protocol for in vitro regeneration of B. senegalensis was developed

Recovery of herbicide-resistant Azuki bean [Vigna angularis (Wild.), Ohwi & Ohashi] plants via Agrobacterium-mediated transformation

Transgenic azuki bean [Vigna angularis (Willd.) Ohwi & Ohashi] plants expressing the hygromycin phosphotransferase (hpt), green fluorescent protein (sgfp) and phosphinothricin acetyltransferase (bar) genes were obtained by Agrobacterium- tumefacients - mediated transformation. A total of 210 epicotyl explants were inoculated with A. tumefaciens strain EHA105, harboring the binary plasmid pZHBG on MS co-cultivation medium supplemented with 100 mM acetosyringone and 10 mg/l of BA. Following selection on MS medium with 15 mg/l of hygromycin, the regenerated adventitious shoots that formed on the induced calli were further screened for sgfp expression before transferred to rooting medium. 31 transgenic plants were obtained with transformation frequency of 14%. The presence of transgenes in transformed azuki bean plants was confirmed by polymerase chain reaction (PCR) and southern blot analysis. Transcription of the bar and hpt genes was assessed by reverse transcription polymerase chain reaction (RT-PCR) analysis. sgfp- positive transgenic plants exhibited functional expression of the bar gene as determined by assaying for resistance to bialaphos applied directly to leaves. This result demonstrates the feasibility of introducing potentially useful agronomic traits into azuki bean through genetic engineering. Key Words: Agrobacterium tumefaciens, bar gene, bialaphos, transgenic, Vigna angulazris. African Journal of Biotechnology Vol.4(1) 2005: 61-6

Dedifferentiation of leaf explants and antileukemia activity of an ethanolic extract of cell cultures of Moringa oleifera

The present study was aimed at developing an efficient protocol for callus induction from the leaves of Moringa oleifera and to investigate its crude extract antileukemia activity on leukemia cells. Several secondary metabolites are present in M. oleifera as the plant serves as reservoirs for various bioactive compounds. Callus cultures of M. oleifera were induced from leaf explants incubated on MS medium supplemented with different concentrations of 2,4-dichloro-phenoxyacetic acid (2,4-D). The crude extracts of the callus were evaluated in vitro for their activity against leukemia cells and hepatocarcinoma. Among the different concentrations, 2,4-D at 0.1 mg/l induced highest frequencies of callus growth index (7.8) when compared with other concentrations. Ethanolic extracts killed about 36% of abnormal cells among primary cells harvested from 3 patients with acute myeloid leukemia (AML) and hepatocarcinoma cells HpG2. These results provide an in vitro evidence and support the traditional use of M. oleifera leaf as a potent source of anticancer. However, more researches are needed at phytochemical and clinical levels to confirm the traditional use of this plant as anticancer.Keywords: Moinga olifera, callus culture, antileukemia, hepatocarcinom

Combining Ability and Heterosis for Yield and Yield Components in Maize

Abstract: The study was conducted at two sites , University of Khartoum the experimental farm, faculty of Agriculture, Shambat and at west of Khartoum state, Elrawkeeb Dry Land Research Station, Sudan, during the summer and winter seasons of 2009 and 2010 respectively. Five inbred lines (2, 3, 6, 277, and 405) were used as lines and two inbred lines namely (66Y and 160) were used as (testers). These lines were crossed together according to line x tester technique to generate 10 F 1 -hybrids, every genotype was planted in rows with 4 m along, 70cm between rows and 25cm between plants. A line × tester method for estimation the general combining ability (GCA) of parent and specific combining ability (SCA) of their F 1 -hybrids was used. Genetic components resulting from additive and nonadditive type of gene action were also estimated. Heterosis was measured as a deviation from the midparents and better-parent. The analysis of variance for combining ability revealed that both GCA and SCA variances were highly significant for most of the studied characters indicating importance of additive as well as non-additive types of gene action in controlling these traits. GCA mean squares for inbred lines were significant (P < 0.01) for all the traits except cob length and number of kernels/row while GCA due to testers was only significant (P<0.05) for 100-kernels weight. Moreover, variances due to SCA were higher in magnitude than GCA for the yield and yield components except cob diameter, number of rows/cob, number of kernels/row and harvest index. GCA to SCA ratios were less than one for most of the traits except cob diameter, number of kernels row/cob, number of kernels/row and harvest index indicating a preponderance of additive over no additive gene action. High positive heterosis for grain yield and its components was found for more than half of the hybrids studied. Crosses involving 160×3 and 66Y×2 produced the highest heterosis. It can be concluded that these parental lines can be desirable parents for hybrids as well as for inclusion in breeding program, since they may contribute favorable alleles in the synthesis of new varietie

Full Length Research Paper - Recovery of herbicide-resistant Azuki bean [Vigna angularis (Wild.), Ohwi & Ohashi] plants via Agrobacterium- mediated transformation

Transgenic azuki bean [Vigna angularis (Willd.) Ohwi & Ohashi] plants expressing the hygromycin phosphotransferase (hpt), green fluorescent protein (sgfp) and phosphinothricin acetyltransferase (bar) genes were obtained by Agrobacterium - tumefacients - mediated transformation. A total of 210 epicotyl explants were inoculated with A. tumefaciens strain EHA105, harboring the binary plasmid pZHBG on MS co-cultivation medium supplemented with 100 mM acetosyringone and 10 mg/l of BA. Following selection on MS medium with 15 mg/l of hygromycin, the regenerated adventitious shoots that formed on the induced calli were further screened for sgfp expression before transferred to rooting medium. 31 transgenic plants were obtained with transformation frequency of 14%. The presence of transgenes in transformed azuki bean plants was confirmed by polymerase chain reaction (PCR) and southern blot analysis. Transcription of the bar and hpt genes was assessed by reverse transcription polymerase chain reaction (RT-PCR) analysis. sgfp- positive transgenic plants exhibited functional expression of the bar gene as determined by assaying for resistance to bialaphos applied directly to leaves. This result demonstrates the feasibility of introducing potentially useful agronomic traits into azuki bean through genetic engineering

Active principle from Moringa oleifera Lam leaves effective against two leukemias and a hepatocarcinoma

Medicinal plants are important elements of indigenous medical system that have persisted in developing countries. Many of the pharmacological principles currently used as anticancer agents were first isolated from plants. However, some important anticancer agents are still extracted from plants because they cannot be synthesized chemically on a commercial scale due to their complex structures that often contain several chiral centers. The aim of this study was to test different extracts from the leaves of Moringa or drumstick tree (Moringa oleifera) for activity against leukemia and hepatocarcinoma cells in vitro. The extracts could kill majority (70 - 86%) of the abnormal cells among primary cells harvested from 10 patients with acute lymphoblastic leukemia (ALL) and 15 with acute myeloid leukemia (AML) as well as a culture of hepatocarcinoma cells (75% death), but most significantly by the hot water and ethanol extracts. In conclusion, M. oleifera may have potential for use as source of natural treatment for diseases such as cancer.Key words: Moringa oleifera, anti-cancer, acute lymphoblastic leukemia, acute myeloid leukemia, hepatocarcinoma

Efficient production of transgenic soybean (Glycine max [L] Merrill) plants mediated via whisker-supersonic (WSS) method

The present study was designed to evaluate the transformation efficiency and proof the capability of whisker supersonic (WSS) method as an alternative option for soybean (Glycine max [L] Merrill) transformation. We compared soybean transformation efficiency obtained by WSS-mediated with that of particle bombardment transformation by carrying out molecular analysis of the T 0 plants in two independent experiments. For this, we used for both transformation techniques the same genotype, the same plasmid and the same selection method. To assess the efficiency of soybean genetic transformation, we evaluated the efficiency of multi gene transformation by the selection with hygromycin and the expression of green fluorescent protein [sGFP (S65T)] resulted from both techniques. Regenerable embryogenic cells were induced from immature cotyledons of soybean c.v Jack on MSD40 media within 3 weeks then proliferated on FN lite liquid media and engineered with pUHG gene construct through both WSS and particle bombardment-mediated transformation. The pUHG was constructed with pUC 19 and contain the hpt gene conferring resistance to hygromycin as a selective marker and sGFP(S65T) as a reporter gene. Fluorescence microscopy screening after the selection of hygromycin, identified the clearly expression of sGFP(S65T) in the transformed soybean embryos. Stable integration of the transgenes was confirmed by polymerase chain reaction (PCR) and Southern blot analysis. The average transformation efficiency achieved with WSS was higher than that obtained by particle bombardment and hence it may represent an alternative method for soybean transformation

Establishment of an in vitro micropropagation protocol for Boscia senegalensis (Pers.) Lam. ex Poir.

This report describes in vitro micropropagation of Boscia senegalensis, so-called famine foods, that helped the people in Darfur and Kordofan, Sudan survive during the 1984–1985 famine. Four types of explants prepared from green mature zygotic embryos were cultured on Murashige and Skoog (MS) medium augmented with 1–5 mg/L 6-benzyladenine (BA). The highest number of shoots per explant (14.3±0.9) was achieved on MS medium supplemented with 3 mg/L BA, while the highest shoot length [(3.5±0.4) cm] was obtained with 1 mg/L BA. The shoot cluster, when subcultured to its same medium, significantly increased the rate of shoot multiplication by the end of the third subculture. The maximum mean number of shoots per explant (86.5±3.6) was produced after three multiplication cycles on 3 mg/L BA-supplemented medium. In vitro induced shoots were excised and rooted on half strength MS medium fortified with 0.25 mg/L indole-3-butyric acid (IBA) to obtain complete plantlets. B. senegalensis-regenerated plantlets obtained in vitro for the first time, were hardened and 95% survived under greenhouse conditions