<p>The synthetic dye industry is a important driving force behind a diverse variety of fields, such as the textile, light, and chemical industries. In particular, the textile industry is a main consumer of these synthetic dyes - 10,000 tons out of the 7 × 107 tons of synthetic dyes produced worldwide are used by textile industries <ahref="https://doi.org/10.1016/j.ecoenv.2021.113160"><sup>[1]</sup></a>.</p>
<p>China is widely regarded as the “world's factory”, and the name holds true for its role in the dye and textile industries. In terms of annual production, export, and retail, China firmly holds first place in the world textile industry. The sector has experienced consistent economic growth over the last decade, now producing nearly 65% of the world’s clothing <ahref="https://doi.org/10.3390/textiles2010010"><sup>[5]</sup></a>. In the dye industry, its fuel production and trade volume rank first in the world (Xu et al., 2002).</p>
<p>However, this economic prosperity has a dark side. The amount of waste water discharged by the textile industry is as high as 900 million tons per year, accounting for the sixth place in industrial discharge <ahref="http://dx.doi.org/10.1088/1755-1315/514/5/052001"><sup>[11]</sup></a>. Untreated textile wastewater contains a high concentration of toxic chemical dyes and heavy metals discharged as effluent, which contaminate local waters, causing serious pollution and posing a serious risk to aquatic ecosystems and human health.</p>
<p>Current treatment methods for industrial wastewater include physical methods, such as filtration, flocculation and adsorption; or chemical methods, like oxidation and ozonation. However, the water-soluble and stable properties of synthetic dyes make them difficult to be removed from wastewater using simple treatment methods <ahref="https://doi.org/10.1016/j.tca.2019.03.024"><sup>[10]</sup></a>, and the possible generation of toxic intermediates raises concerns on ecofriendliness. These methods also require expensive machinery and chemicals, which are unaffordable for many small scale factories and companies <ahref="https://doi.org/10.1016/j.jece.2020.105012"><sup>[10]</sup></a>.</p>
<p>However, this economic prosperity has a dark side. The amount of waste water discharged by the textile industry is as high as 900 million tons per year, accounting for the sixth place in industrial discharge <ahref="http://dx.doi.org/10.1088/1755-1315/514/5/052001"><sup>[12]</sup></a>. Untreated textile wastewater contains a high concentration of toxic chemical dyes and heavy metals discharged as effluent, which contaminate local waters, causing serious pollution and posing a serious risk to aquatic ecosystems and human health.</p>
<p>Current treatment methods for industrial wastewater include physical methods, such as filtration, flocculation and adsorption; or chemical methods, like oxidation and ozonation. However, the water-soluble and stable properties of synthetic dyes make them difficult to be removed from wastewater using simple treatment methods <ahref="https://doi.org/10.1016/j.tca.2019.03.024"><sup>[11]</sup></a>, and the possible generation of toxic intermediates raises concerns on ecofriendliness. These methods also require expensive machinery and chemicals, which are unaffordable for many small scale factories and companies <ahref="https://doi.org/10.1016/j.jece.2020.105012"><sup>[11]</sup></a>.</p>
<figcaption>Current treatment methods for industrial wastewater</figcaption>
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<h2>Project</h2>
<hr>
<h5>Methylene blue (MB)</h5>
<p>Is a thiazine dye under polymethine dyes. It is the most common and highest-consumed dye in the textile industry, and is considered one of the most popular clothing colourants <ahref="https://doi.org/10.3390/w14020242"><sup>[11]</sup></a>. Worryingly, it is toxic, carcinogenic, mutagenic, and can be fatal to humans at concentrations over 5 mk/kg <ahref="https://doi.org/10.1016/j.rineng.2022.100678"><sup>[10]</sup></a>. It is also water-soluble and forms a highly coloured sub-product in water, which decreases sunlight transmittance and oxygen solubility in aquatic environments. This affects the photosynthetic and respiratory activity of aquatic life, and has dire impacts on biodiversity. It is difficult to degrade by traditional means as it is thermal and light-stable.</p>
<p>Is a thiazine dye under polymethine dyes. It is the most common and highest-consumed dye in the textile industry, and is considered one of the most popular clothing colourants <ahref="https://doi.org/10.3390/w14020242"><sup>[12]</sup></a>. Worryingly, it is toxic, carcinogenic, mutagenic, and can be fatal to humans at concentrations over 5 mk/kg <ahref="https://doi.org/10.1016/j.rineng.2022.100678"><sup>[11]</sup></a>. It is also water-soluble and forms a highly coloured sub-product in water, which decreases sunlight transmittance and oxygen solubility in aquatic environments. This affects the photosynthetic and respiratory activity of aquatic life, and has dire impacts on biodiversity. It is difficult to degrade by traditional means as it is thermal and light-stable.</p>
<h5><i>S. elongatus</i></h5>
<p><i>S. elongatus</i> is a species of cyanobacteria, which have long been considered promising candidates for microbial cell factories. They rely on a cost-effective minimal growth requirement; CO2, light, water, and minimal nutrients. Their photosynthetic nature also makes them an attractive green option for the atmospheric reduction of CO2 on top of biosynthesis of the proteins of interest. S. elongatus also possesses a high salt and metal tolerance, allowing them to be the optimum chassis for microbial remediation. We hope to improve its bioremediative ability by engineering it to express homologous ligninolytic isozymes characterized in other species.</p>
<h5>Enzymes</h5>
<p>Laccases (Lac) belong to the enzyme family of multi-copper oxidases (MCOs) and can be obtained from a wide variety of species such as bacteria <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[11]</sup></a>, brown and white-rot fungi (Heinzkill et al., 1998), and various plant and algae species <ahref="https://doi.org/10.1186/s12934-019-1248-0"><sup>[2]</sup></a>. Together with lignin peroxidase (LiP) and manganese peroxidase (MnP), this enzyme combination has displayed the ability to catalyze the breakdown of various inorganic and organic substances such as wood, plastic, paint, and jet fuel into nutrients in many species (Viswanath et al., 2014), and have been proven to be the main enzymes responsible for catalyzing the degradation of MB <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[11]</sup></a>.</p>
<p>Laccases (Lac) belong to the enzyme family of multi-copper oxidases (MCOs) and can be obtained from a wide variety of species such as bacteria <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[12]</sup></a>, brown and white-rot fungi (Heinzkill et al., 1998), and various plant and algae species <ahref="https://doi.org/10.1186/s12934-019-1248-0"><sup>[2]</sup></a>. Together with lignin peroxidase (LiP) and manganese peroxidase (MnP), this enzyme combination has displayed the ability to catalyze the breakdown of various inorganic and organic substances such as wood, plastic, paint, and jet fuel into nutrients in many species (Viswanath et al., 2014), and have been proven to be the main enzymes responsible for catalyzing the degradation of MB <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[12]</sup></a>.</p>
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@@ -58,8 +58,8 @@
<divclass="col-lg-12">
<h2>System</h2>
<hr>
<p>We aim to design plasmids with inserts of homologous variants on these three genes from species such as <i>B. thuringiensis</i><ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[11]</sup></a> and <i>T. pubescens</i><ahref="https://www.cabdirect.org/cabdirect/abstract/20183369455"><sup>[14]</sup></a> . The plasmid will first be transformed into DH5a E. coli for cloning, then electroporation for homologous recombination-based genomic insertion will be performed based on a protocol by Omayra et al. (2022) into the genome of S. elongatus. Expression of our vectors will be confirmed by PCR and MB-degrading activity will be quantified by spectrophotometric analysis. </p>
<p>MB degraded metabolites' toxicity has been found to be lower than that of the parent compound <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[11]</sup></a>. To investigate the toxicity of the degradation products, which have been proposed to be aromatic amines, raw and treated EC50 values will be used for comparison</p>
<p>We aim to design plasmids with inserts of homologous variants on these three genes from species such as <i>B. thuringiensis</i><ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[12]</sup></a> and <i>T. pubescens</i><ahref="https://www.cabdirect.org/cabdirect/abstract/20183369455"><sup>[14]</sup></a> . The plasmid will first be transformed into DH5a E. coli for cloning, then electroporation for homologous recombination-based genomic insertion will be performed based on a protocol by Omayra et al. (2022) into the genome of S. elongatus. Expression of our vectors will be confirmed by PCR and MB-degrading activity will be quantified by spectrophotometric analysis. </p>
<p>MB degraded metabolites' toxicity has been found to be lower than that of the parent compound <ahref="https://doi.org/10.1016/j.enzmictec.2022.109999"><sup>[12]</sup></a>. To investigate the toxicity of the degradation products, which have been proposed to be aromatic amines, raw and treated EC50 values will be used for comparison</p>
</div>
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<ul>[6] Khan, I., Saeed, K., Zekker, I., Zhang, B., Hendi, A. H., Ahmad, A., ... & Khan, I. (2022). Review on methylene blue: its properties, uses, toxicity and photodegradation. Water, 14(2), 242. https://doi.org/10.3390/w14020242</ul>
<ul>[7] Kishor, R., Purchase, D., Saratale, G. D., Saratale, R. G., Ferreira, L. F. R., Bilal, M., ... & Bharagava, R. N. (2021). Ecotoxicological and health concerns of persistent coloring pollutants of textile industry wastewater and treatment approaches for environmental safety. Journal of Environmental Chemical Engineering, 9(2), 105012. https://doi.org/10.1016/j.jece.2020.105012</ul>
<ul>[8] Liu, Q. (2020). Pollution and treatment of dye waste-water. In IOP Conference Series: Earth and Environmental Science (Vol. 514, No. 5, p. 052001). IOP Publishing. http://dx.doi.org/10.1088/1755-1315/514/5/052001</ul>
<ul></ul>
<ul>[9] Oladoye, P. O., Ajiboye, T. O., Omotola, E. O., & Oyewola, O. J. (2022). Methylene blue dye: Toxicity and potential elimination technology from wastewater. Results Eng 16: 100678. https://doi.org/10.1016/j.rineng.2022.100678</ul>
<ul>[10] Salimi, A., & Roosta, A. (2019). Experimental solubility and thermodynamic aspects of methylene blue in different solvents. Thermochimica Acta, 675, 134-139. https://doi.org/10.1016/j.tca.2019.03.024</ul>
<ul>[11] Wu, K., Shi, M., Pan, X., Zhang, J., Zhang, X., Shen, T., & Tian, Y. (2022). Decolourization and biodegradation of methylene blue dye by a ligninolytic enzyme-producing Bacillus thuringiensis: degradation products and pathway. Enzyme and Microbial Technology, 156, 109999. https://doi.org/10.1016/j.enzmictec.2022.109999</ul>
<ul>[12] Xu W. (2022). Current Situation and Prospect of waste-water Treatment in Dye Industry [J]. Dye Industry, 39(6): 35- 39.</ul>
<ul>[9] Ming Xu, Yuanyuan Cui, Meng Hu, Xinkai Xu, Zhechi Zhang, Sai Liang, Shen Qu. (2019). Supply chain sustainability risk and assessment https://doi.org/10.1016/j.jclepro.2019.03.307</ul>
<ul>[10] Oladoye, P. O., Ajiboye, T. O., Omotola, E. O., & Oyewola, O. J. (2022). Methylene blue dye: Toxicity and potential elimination technology from wastewater. Results Eng 16: 100678. https://doi.org/10.1016/j.rineng.2022.100678</ul>
<ul>[11] Salimi, A., & Roosta, A. (2019). Experimental solubility and thermodynamic aspects of methylene blue in different solvents. Thermochimica Acta, 675, 134-139. https://doi.org/10.1016/j.tca.2019.03.024</ul>
<ul>[12] Wu, K., Shi, M., Pan, X., Zhang, J., Zhang, X., Shen, T., & Tian, Y. (2022). Decolourization and biodegradation of methylene blue dye by a ligninolytic enzyme-producing Bacillus thuringiensis: degradation products and pathway. Enzyme and Microbial Technology, 156, 109999. https://doi.org/10.1016/j.enzmictec.2022.109999</ul>
<ul>[13] Xu W. (2022). Current Situation and Prospect of waste-water Treatment in Dye Industry [J]. Dye Industry, 39(6): 35- 39.</ul>
<ul>[14] Zhang, T., Zhang, B., & Li, Y. (2018). Decolorization of thiazine dye methylene blue by three white-rot fungi. Mycosystema, 37(9), 1243-1250 https://www.cabdirect.org/cabdirect/abstract/20183369455</ul>
