Validation of a genus-specific gene; TPS, used as internal control in quantitative Real Time PCR of transgenic cotton

Abstract Identification of genes with invariant levels of gene expression is a prerequisite for validating transcriptomic changes accompanying development. Ideally expression of these genes should be independent of the morphogenetic process or environmental condition.We report here the validation of internal control gene i.e.TPS (trehalose 6-phosphate-synthase) in cotton (Gossypium spp), using TaqMan system in quantitative Real Time PCR (qRT-PCR). The Gene expression was tested in five different G. hirsutum cultivars including Coker 312, Acala SJ, ZETA 2, Taghva, Neishabour and a diploid wild type; G. barbadense. Identical amplicons were obtained within these cultivars. No amplifications was achieved when DNA samples from barley (Hordeum vulgare), maize (Zea mays), rice (Oryza sativa)

Transgenic crops with an improved resistance to biotic stresses. A review

Introduction. Pests, diseases and weeds (biotic stresses) are significant limiting factors for crop yield and production. However, the limitations associated with conventional breeding methods necessitated the development of alternative methods for improving new varieties with higher resistance to biotic stresses. Molecular techniques have developed applicable methods for genetic transformation of a wide range of plants. Genetic engineering approach has been demonstrated to provide enormous options for the selection of the resistance genes from different sources to introduce them into plants to provide resistance against different biotic stresses. Literature. In this review, we focus on strategies to achieve the above mentioned objectives including expression of insecticidal, antifungal, antibacterial, antiviral resistance and herbicide detoxification for herbicide resistance. Conclusion. Regardless of the concerns about commercialization of products from genetically modified (GM) crops resistant to biotic stresses, it is observed that the cultivation area of these crops is growing fast each year. Considering this trend, it is expected that production and commercialization of GM crops resistant to biotic stresses will continue to increase but will also extend to production of crops resistant to abiotic stresses (e.g. drought, salinity, etc.) in a near future

Effect of estradiol on miR-21& miR-155 expression in promyelocytic leukemia-derived cell line NB4

Background: Due to the estrogen participation in modulating the proliferation and commitment of stem cells and the effects of miR-21 and miR-155 expression on reduced proliferation and colony formation of acute myeloid leukemia (AML), the aim of the present study was to evaluate the effect of estradiol on expression of miR-21 and miR-155 in the NB4 cell line, as an acute promyelocytic leukemia (APL). Materials and Methods: In the present experiment, NB4 cells were treated with different quantities of estradiol (5, 25, 50, 75, 100, 150, 200, 250 μg/ml) and vehicle control for 24 and 48 hours. Viability, apoptosis, and cellular proliferation were estimated by trypan blue exclusion, flow cytometry, and MTT assays, respectively. The level of miR-155 and miR-21 expression was studied using absolute quantitative real-time PCR. Results: Results showed that estradiol in the effective dose (200 μg/ml) led to decreased cellular viability (in a dose dependent manner, P = 0.004) and apoptosis of NB4 cells. In addition, the expressions of miR-155 and miR-21 were significantly and dose-dependently decreased (p<0.05). Conclusion: Estradiol at the effective dose caused apoptosis in NB4 cell line. This substance can be used as a drug for the treatment of APL. However, further assessments are needed to support the effectiveness of estradiol in the treatment of APL. © 2020 Shahid Sadoughi University of Medical Sciences

Enhanced insect resistance to bollworm (Helicoverpa armigera) in cotton containing a synthetic cry1Ab gene

72-77In order to investigate stability of the cry1Ab gene and resistance in the T1 generation of transgenic cotton (Gossypium hirsutum var. Coker 100), transgenic seeds were analyzed by PCR, Southern and Western blotting and bioassayed. Seed samples from transgenic lines (T0) along with seeds from non-transgenic lines s as control were used. PCR analysis showed the presence of cry1Ab gene in 63 out of the 150 T1 plants. Integration of the cry1Ab gene into the genome of transgenic plants was confirmed by Southern blotting. Western immunoblot analysis of transgenic plants revealed the presence of a band with MW of 67 kDa using anti- Cry1Ab-polyclonal antiserum. Bioassay tests indicated that the Cry1Ab protein was active and that the larvae died in the first days. Significant differences were observed in the mortality rate, growth rate and extent of damage on the leaves between the transgenic and non-transgenic plants. The average mortality of the larvae in line 17 was 100%, in comparison with the control. These results indicated the relative advantage of line 17, which carries one copy of the cry1Ab gene in comparison with the other lines

Interaction between Bt-transgenic cotton and the whitefly’s parasitoid, Encarsia formosa (Hymenoptera: Aphelinidae)

Transgenic Bt cotton developed against lepidopteran pests may not be compatible with parasitoid of secondary pests such as Bemisia tabaci which attack many plants such as cotton. In this study, the effects of Bt cotton on the demographic parameters of Encarsia formosa, parasitoid of B. tabaci were assessed. The data were analysed using the age specific, two-sex life table parameters. The results indicated that pre-adult developmental time, the total preoviposition period (TPOP) and the adult preoviposition period (APOP) in the Bt cotton were significantly longer than in the non-Bt cotton. Also, fecundity and body size in both lines were significantly different. The fecundity was 23.64±0.73 and 43.75±0.89 eggs/females in the Bt and non-Bt cotton, respectively. All the population parameters were affected by the Bt cotton. The intrinsic rate of increase (r) was 0.15 day-1 in the non-Bt cotton but it was 0.10 day-1 in the Bt cotton. The finite rate of increase (λ) was 1.11 day-1 in the non-Bt cotton whilst it was 1.08 in the Bt cotton. The net reproductive rate (R0) in the non-Bt cotton was 36.75 but in the Bt cotton these parameters showed 19.62 offspring/individual. Also, the mean generation time (T ) in the non-Bt and Bt cotton was 22.69 and 27.79 days, respectively. The results illustrated, that although transgenic crops are effective tools for management of the target pests, they can adversely affect, either directly or indirectly, the natural enemies dependent on these plants

Agrobacterium Mediated Transformation of Somatic Embryos of Persian Walnut Using fld Gene for Osmotic Stress Tolerance

Somatic embryos of Persian walnut were transformed with two strains of Agrobacterium tumefaciens i.e. LBA4404 and C58, and two plasmids, namely, pBI121 with nptII and gus genes for improving the transformation protocol, and p6u-ubi-FVTI plasmid containing the hpt and fld genes. The transformation frequency was 10%. PCR and RT-PCR analysis proved the presence and expression of the genes. The transgenic and non-transgenic somatic embryos of Persian walnut were exposed to four salinity levels (0, 50, 100, and 200 mM NaCl) and four osmotic stress (0, 1.5, 5, and 10% PEG) levels. After 20 days, the number of survived, secondary and cotyledonary somatic embryos, as well as fresh and dry weights of embryos were evaluated. In addition, the transgenic and non-transgenic regenerated plantlets with 3 leaves and 2.5 cm length were subjected to 200 mM NaCl. In both experiments, the main effects of fld-transformation and stress treatments on evaluated parameters were significant. Transgenic somatic embryos showed no significant differences at 0 and 200 mM NaCl and 0 and 1.5% PEG. Significant differences of transgenic vs. non-transgenic somatic embryos were observed at 50 and 100 mM NaCl and 5 and 10% PEG. Non-transgenic plantlets on medium containing 200 mM NaCl showed complete necrosis and died after 10 days, while transgenic lines continued growth until 45 days. Our results clearly showed that expression of fld gene increased stress tolerance in fld transformant lines of walnut, and also revealed that expression of this specific cyanobacterial protein may provide a powerful tool to improve tolerance to environmental stresses.Fil: Sheikh Beig Goharrizi, M. A.. Universidad de Teherán; IránFil: Dejahang, A.. Universidad de Teherán; IránFil: Tohidfar, M.. Shahid Beheshti University; IránFil: Izadi Darbandi, A.. Universidad de Teherán; IránFil: Carrillo, Nestor Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Hajirezaei, M. R.. Leibniz Institute of Plant Genetics and Crop Plant Research; AlemaniaFil: Vahdati, K.. Universidad de Teherán; Irá

An introduction to global production trends and uses, history and evolution, and genetic and biotechnological improvements in cotton

Cotton plant has been domesticated in tropical and sub‐tropical climates of the world but severe climate is not suitable for good lint yields. Globally, cotton is grown on an area of more than 30 million hectare and possesses a global production of more than 70 million tons of seed cotton. The average yield of seed cotton in the world is more than 2000 kg/ha and the largest cotton producer in the world (China) is getting an average yield that is double to the world average. China, India, the United States, and Pakistan are the largest seed cotton producers in the world. Cotton crop possesses an inevitable role in the global industries, economy, and culture. In this chapter, the history and evolution of the cotton, the global production trends of cotton, uses, and the role of biotechnology in improving cotton production have been discussed

Genetic transformation of a hepatoprotective plant, Phyllanthus amarus

Phyllanthus amarus Schum & Thonn. is a source of various pharmacologically active compounds such as phyllanthin, hypophyllanthin, gallic acid, catechin, and nirurin, a flavone glycoside. A genetic transformation method using Agrobacterium tumefaciens was developed for this plant species for the first time. Shoot tips of full grown plants were used as explants for Agrobacteriummediated transformation. Transgenic plants were obtained by co-cultivation of shoot tips explants and A. tumefaciens strain LBA4404 containing the pCAMBIA 2301 plasmid harboring neomycin phosphotransferase II (NPT II) and β- glucuronidase encoding (GUS) genes in the T-DNA region in the presence of 200 μM acetosyringone. Integration of the NPT II gene into the genome of transgenic plants was verified by PCR and Southern blot analyses. Expression of the NPT II gene was confirmed by RT-PCR analysis. An average of 25 explants was used, out of which an average of 19 explants produced kanamycin-resistant shoots, which rooted to produce 13 complete transgenic plants