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Plant Biotechnology
Published in Firdos Alam Khan, Biotechnology Fundamentals, 2020
The Agrobacterium-mediated technique involves the natural gene transfer system in the bacterial plant pathogens of the genus Agrobacterium. In nature, Agrobacterium tumefaciens and Agrobacterium rhizogenes are the causative agents of the crown gall and the hairy root diseases, respectively. The utility of Agrobacterium as a gene transfer system was first recognized when it was demonstrated that these plant diseases were produced because of the transfer and integration of genes from the bacteria into the genome of the plant. Both Agrobacterium species carry a large plasmid (small circular DNA molecule) called Ti in A. tumefaciens and Ri in A. rhizogenes. A segment of this plasmid designated T-(for transfer) DNA is transmitted by this organism into individual plant cells, usually within wounded tissue. The T-DNA segment penetrates the plant cell nucleus and integrates randomly into the genome where it is stably incorporated and inherited like any other plant gene in a predictable, dominant Mendelian fashion. Expression of the natural genes on the T-DNA results in the synthesis of gene products that direct the observed morphological changes such as tumor or hairy root formation. In genetic engineering, the tumor-inducing genes within the T-DNA, which cause the plant disease, are removed and replaced by foreign genes. These genes are then stably integrated into the genome of the plant after infection with the altered strain of Agrobacterium, just like the natural T-DNA. Because all tumor-inducing genes are removed, the gene transfer does not induce any disease symptoms. This reliable method of gene transfer is well suited for plants that are susceptible to infection by Agrobacterium. Unfortunately, many species, especially economically important legumes and monocotyledons such as cereals, do not respond positively to Agrobacterium-mediated transformation (Figure 6.6).
Agricultural biotechnology
Published in Firdos Alam Khan, Biotechnology Fundamentals, 2018
The Agrobacterium-mediated technique involves the natural gene transfer system in the bacterial plant pathogens of the genus Agrobacterium. In nature, Agrobacterium tumefaciens and Agrobacterium rhizogenes are the causative agents of the crown gall and the hairy root diseases, respectively. The utility of Agrobacterium as a gene transfer system was first recognized when it was demonstrated that these plant diseases were actually produced as a result of the transfer and integration of genes from the bacteria into the genome of the plant. Both Agrobacterium species carry a large plasmid (small circular DNA molecule) called Ti in A. tumefaciens and Ri in A. rhizogenes. A segment of this plasmid designated T-(for transfer) DNA is transmitted by this organism into individual plant cells, usually within wounded tissue. The T-DNA segment penetrates the plant cell nucleus and integrates randomly into the genome where it is stably incorporated and inherited like any other plant gene in a predictable, dominant Mendelian fashion. Expression of the natural genes on the T-DNA results in the synthesis of gene products that direct the observed morphological changes such as tumor or hairy root formation. In genetic engineering, the tumor-inducing genes within the T-DNA, which cause the plant disease, are removed and replaced by foreign genes. These genes are then stably integrated into the genome of the plant after infection with the altered strain of Agrobacterium, just like the natural T-DNA. Because all tumor-inducing genes are removed, the gene transfer does not induce any disease symptoms. This reliable method of gene transfer is well suited for plants that are susceptible to infection by Agrobacterium. Unfortunately, many species, especially economically important legumes and monocotyledons such as cereals, do not respond positively to Agrobacterium-mediated transformation (Figure 6.6).
Microalgae based wastewater treatment: a shifting paradigm for the developing nations
Published in International Journal of Phytoremediation, 2021
Nandini Moondra, Namrata D. Jariwala, Robin A. Christian
The pH variation observed in the effluent was due to photosynthetic CO2 assimilation (Daliry et al. 2017; Shahid et al. 2020) and nitrification (Schumacher and Sekoulov 2003). High pH helped in the removal of suspended solids through auto-flocculation, and also in phosphates reduction via precipitation (Hoffmann 1998). Cellular uptake (Singh and Pandey 2018), assimilation (Wang et al. 2017) and photosynthesis (Cai et al. 2013) also contributed to phosphate removal. Phosphates are majorly used for energy transfer, DNA and RNA generation in the algal cell (Hwang et al. 2016). Ammonia was reduced in the study due to the active transport and direct utilization by microalgal cells (Han et al. 2019), nitrification and ammonia stripping (Lv et al. 2017). Nitrogen uptake resulted in the formation of peptides, proteins, vitamins, enzymes, chlorophyll, energy transfer molecules (ATP, ADP) and genetic material (Han et al. 2019). COD removal was slightly lesser than nutrients, as an increase in nitrate concentration caused interference in COD removal (Sawyer et al. 2003). COD removal was mainly due to sufficient DO concentration, photosynthetic effect (Wang et al. 2010) and low F/M ratio (Choi and Lee 2012). F/M ratio in the case of algal treatment ranges between 0.05 and 0.1 (Choi and Lee 2012), which is similar to an ASP system’s extended aeration process. A low F/M ratio also leads to less sludge generation as the microorganisms in the system are in the endogenous phase (Metcalf and Eddy 2003).
Identification of a novel strain of fungus Kalmusia italica from untouched marine soil and its heavy metal tolerance activity
Published in Bioremediation Journal, 2021
S. Sumathi, V. Priyanka, V. Krishnapriya, K. Suganya
Transfer of fresh cells (150 mg) to a 1.5-ml sterilized Eppendorf tube containing 300 μl of TES extraction buffer (0.2 M Tris-HCl at pH 8, 10 mM EDTA at pH 8, 0.5 M NaCl, 1% SDS) and acid-washed, sterilized sea sand or 0.5 mm glass beads. Keep for 2 min with such a handheld disposable homogenizer fitting the 1.5-ml microcentrifuge tube. Vortex extracts for 30 s and added 200 μl proteinase. Vortex TES extraction buffer and completely mix and position tubes at 65 °C for 30 min in a water bath. Add 7.5 M ammonium acetate (250 μl) by 1⁄2 volume. In the refrigerator, mix and incubate the samples over ice or 5 °C for 10 min. Centrifuge the samples for 15 min. Transfer the supernatant to one of a new tube and add an equal volume of ice-cold isopropanol (500 microns). Incubate the tubes for 1–2 h at 20 °C. Centrifuge for 10 min, decant the supernatant and wash DNA pellet with cold 70% ethanol at 800 μl. On clean sterile paper towels, turn upside down tubes to air-dry DNA for 10–15 min. Twice-repeated extractions of Elute DNA from the pellet with 250 μl of 1X TE buffer. Transfer DNA solution to a 1.5 ml eppendorf tube, add RNase A 5 μl (20 mg/ml), and incubate for 60 min at 37 °C. The dry DNA pellet may be shipped for analysis to other laboratories or dissolved in a buffer of 100 μl l × TE.