Difference between revisions of "Team:UESTC-China/description"

 
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<p><img src="https://static.igem.org/mediawiki/2017/1/10/T--UESTC-China--xianweijing.png" alt="" class="hhh" /></p>
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<a href="#Overview">Overview</a>
 
<a href="#Overview">Overview</a>
 
</li>
 
</li>
<li>
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<li> <a href="#Harm">Distributions of TCP</a> </li>
<a href="#Construct-the-pathway" style="font-size: 14.5px;">Construct the pathway</a>
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<li> <a href="#Technology">Ways to solve </a> </li>
</li>
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<li> <a href="#Strategy">Our super tobacco</a> </li>
<li>
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<a href="#Improve-the-function">Improve the function</a>
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</li>
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<li>
 
<li>
 
<a href="#References">References</a>
 
<a href="#References">References</a>
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<div id="Overview">
 
<div id="Overview">
 
<h2>Overview</h2>
 
<h2>Overview</h2>
 
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<p>The progress of the chemical industry and agriculture has brought great convenience to our lives. But a large number of chemical pollutants which were toxic and difficult to degrade have been discharged to the rivers and soils, seriously polluting our environment. Xenobiotic organohalognse played a large part among all the chemical pollutants. After collecting a questionnaire for industrial and agricultural producers and doing a soil research in most areas of China, our team aimed at degrading an organic chloride, 1,2,3-Trichloropropane (1,2,3-TCP), which was less concerned by environmental protection department. In this summer, we decided to use synthetic biology to achieve plant degradation of 1,2,3-Trichloropropane by transferring three enzymes into tobacco which is environmentally friendly and the product, glycerol, is recyclable in tobacco.</p>
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/1/1a/T--UESTC-China--design_1.jpg" style="width: 95%;" /></p>
+
<p>As early as 2013, for the degradation of TCP, 2013iGEM team of our school had built the plasmid of DhaA31 and HheC-W249P , and used E. coli as a chassis biology where the two enzymes achieved the purpose of degradation. Subsequent studies have shown that the triple enzyme cascade pathway consisting of DhaA31, HheC and EchA can convert TCP into non-toxic glycerol[1]. In order to increase degradation efficiency, we used the excellent mutant DhaA31[2], HheC-W249P[3], together with the wild type EchA to realize gene stacking. We used synthetic biological means to construct a plasmid carried three genes firstly. Compared to physical and microbial degradation, phytoremediation is more sustained and efficient by which we can achieve the degradation pathway. Considering that the plant is likely to express EchA under stress, we will build a plasmid containing only the first two genes, and finally get a highly efficient phytoremediation pathway of TCP. In the process of degradation, toxic effects have decreased as well as the growth cycle of plants has extended.</p>
+
 
</div>
 
</div>
  
<div id="Construct-the-pathway">
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<div id="Harm">
<h2>Construct the pathway of phytoremediation</h2>
+
<h2>The distributions of pollutant 1,2,3-TCP</h2>
 
+
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/c/cc/T--UESTC-China--design_2.png" style="width: 75%;" /></p>
+
  <p>1,2,3-TCP is an emerging organic pollutant which is usually been used as industrial solvents[1] and raw materials for producing 1,1,2,3-Tetrachloropropene or other chemicals substances[2]. In agriculture, it has been used as one of the ingredients of soil fumigants and it is also a harmful byproduct of other pesticides. The global production of 1,2,3-TCP reached about 50,000 tons annually[3]. Because 1,2,3-TCP is hard to degrade naturally in the environment, it would cause a huge damage to the groundwater and the soil if discharged casually.</p>
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/d/d1/T--UESTC-China--design_3.jpg" style="width: 100%;" /></p>
+
                                          <p>One of the most serious pollution of 1,2,3-TCP was discovered in California. It has spread to all over the California since the 1940s when Dow Chemical and Shell started selling two soil fumigants (D-D and Telone) which include 1,2,3-TCP. Although 1,2,3-TCP was banned to use as soil fumigants in the 1990s, there was still a large amount of 1,2,3-TCP remained, to be a threat to the environment and people's lives seriously[4].</p>
<p>DhaA, HheC, EchA these three enzymes are the key enzymes to degrade TCP and produce glycerol. Their expression activity in tobacco determines the degradation efficiency of TCP. First, we selected pCaMV35s, a commonly used promoters in plant and the application of 2A peptide strategy to achieve the stable expression of three enzymes in tobacco[4]. In order to realize the degradation of TCP and realize the production of beneficial glycerol, we constructed a plasmid model containing three enzymes of DhaA31, HheC-W249P and EchA. When DhaA31 works independently in tobacco, 1,2,3-TCP can be degraded to 2,3-DCP. When HheC-W249P works independently in tobacco, 2,3-DCP can be degraded to ECH, and CPD is degraded to GDL, when EchA work independently in tobacco, it can degrade ECH into CPD and degrade GDL into glycerol. In 2013, UESTC-China induced DhaA31and HheC-W249P in E.coli, and this year we induced the two enzymes together with EchA in tobacco. In this way, 1,2,3-TCP can be degraded into glycerol, so we first designed a three-gene plasmid. Considering the plant itself may exist EchA[5], in order to avoid too many exogenous enzymes and affect plant metabolism, so we designed a plasmid with only the first two enzymes.</p>
+
                                       
 +
  <p class="pic"><img src="https://static.igem.org/mediawiki/2017/7/7c/T--UESTC-China--description_1.jpg" style="width: 55%;"/></p>
 +
  <p class="mid">Figure 1. Water systems where significant levels of the 1,2,3-TCP have been detected in California</p>
 +
<p>Dr. Yong Qian from University of Geosciences of China has explored the behavior of 1,2,3-TCP in the groundwater. He found 1,2,3-TCP still huge concentration (3890mg / L) underground in an abandoned factory that was only run from 1976 to 1979 in 2016 [5], which showed the great stability of 1,2,3-TCP in the groundwater and soil (Fig. 2). </p>
 +
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/8/8b/T--UESTC-China--222.png" style="width: 60%;"/></p>
 +
  <p class="mid">Figure 2. The distribution of underground 1,2,3-TCP pollution in this factory [5].</p>
 +
<p>Meanwhile, some studies showed that the adhesion coefficient of 1,2,3-TCP is very low[5]. It means that 1,2,3-TCP can be easily diffused into our daily use water. Its potential carcinogenicity and damage to kidney will threaten the health of human beings. In the recent past 10 years, more and more tests showing the existence of 1,2,3-TCP among worldwide drinking water is a sound proof (Fig. 3). </p>
 +
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/4/4b/T--UESTC-China--description_3.png" style="width: 90%;"/></p>
 +
  <p class="mid">Figure 3. 1,2,3-TCP has been detected in hundreds of surface water and drinking water sources </p>
 +
<p>Based on the above information, 1,2,3-TCP is absolutely a threatening pollutant. However, attentions are far more than enough to be paid on 1,2,3-TCP; and a lot of countries have not put 1,2,3-TCP as the test list of water quality. Since the current situation has been known to us, our team hopes to find a feasible way to stop 1,2,3-TCP by attracting the attentions of the whole society through doing advertisement and iGEM competition before it might cause great damage to the health of human beings. </p>
 
</div>
 
</div>
  
<div id="Improve-the-function">
+
<div id="Technology">
<h2>Improve the function of our tobacco</h2>
+
  <h2>Ways to solve the pollution of 1,2,3-TCP</h2>
 
+
  <p>Then the question is how to solve the problem of 1,2,3-TCP pollution? Traditional methods to deal with 1,2,3-TCP includes granular activated carbon (GAC)、hydrogen release compound (HRC)、reductive dechlorination by zero valent iron(ZVI) and son on[6]~[8]. However, some of these methods are either too inefficient or too expensive to be used within natural conditions (Fig. 4). </p>
<p>Only degrading 1,2,3-TCP is not enough, we also pursue a more efficient way. Considering that the three enzymes expressed mainly in the roots where tobacco are in direct contact with wastewater or soil containing 1,2,3-TCP, we designed the plant root-specific expression promoter pYK10 helping enzymes fixed at the root[6]. Thus the development of super-tobacco roots is particularly important, in order to improve the biomass of roots to increase the yield of enzyme, the CKX gene is selected by us[7]. Finally, in order to make the three enzymes more susceptible to TCP, we chose the cell wall localization signal peptide AO-S to achieve fusion expression with three enzymes (at the same time AO-S also has the role of stabling enzyme expression)[8]. Through the improvements above, we get more powerful super tobacco at last.</p>
+
  <p class="pic"><img src="https://static.igem.org/mediawiki/2017/3/34/T--UESTC-China--description_4.jpg" style="width: 95%;"/></p>
<p>We plan to apply super-tobacco to real life in the future, and for this purpose we should take full account of biosafety and maneuverability. We will induce AdCP gene into our plans in the future because of its capability to lead to pollen abortion. At the same time, chloroplast transformation will be taken into consideration to avoid gene flow and improve gene expression. Thus, we can both meet the actual needs and ensure the biosafety. </p>
+
  <p class="mid">Figure 4. Treatability tests with 1,2,3-TCP-contaminated groundwater/soil</p>
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/a/ac/T--UESTC-China--design_4.jpg" style="width: 90%;" /></p>
+
  <p>Since 1,2,3-TCP can hardly be dealt with traditional ways, Microbial remediation of 1,2,3-TCP is getting praised. Early studies have shown that 1,2,3-TCP can be converted into CO<sub>2</sub>、H<sub>2</sub>O and HCl by biological catalysis through Oxidation of metabolic mechanism with O<sub>2</sub>. So people are trying to find degradation method by using aerobe. But until now, all the tests to gather and filtrate aerobe to degrade 1,2,3-TCP have failed. However, scientists have found a few categories of bacteria that can degrade 1,2,3-TCP in absolute anaerobic environment. However, due to its strict requirement, the conversion efficiency is not high enough to get popular. In recent years, some studies have started to apply genetic engineering method to inject some enzymes in the seek of 1,2,3-TCP degradation. Some good results have been obtained but with some limitations. Firstly, these bacteria require very strict nutrition requirement and there only exits weak competition among them. Secondly, the diffusion of antibiotic resistance gene can be easily triggered. Finally, this method rely heavily on some specific induction condition[9]. We are not satisfied with these disadvantages and thus we hope to find a 1,2,3-TCP degradation method that is energy-efficient and sustainable. The burgeoning “Phytoremediation” has come into our attention. </p>
 
</div>
 
</div>
  
<div id="References">
+
<div id="Strategy">
<h2>References</h2>
+
  <h2>Our super tobacco</h2>
 
+
  <p>As one of the new “Green remediation” strategy,phytoremediation shows enormous potential. Compared to microbial remediation, the most important advantage of phytoremediation is that phytoremediation comes with the potential to dispose pollutants by small amount of nutrition input. This is due to its unique feature of photosynthetic autotrophs system. By doing pollutant disposition, at the same time, plants can also help stabilize the soil, purify the water and clear air pollution[10]~[11]. There are four different ways of “Phytoremediation”: phytoextraction、phytostabilization、phytovolatilization and phytodegradation (Fig. 5). </p>
<ol>
+
<p class="pic"><img src="https://static.igem.org/mediawiki/2017/f/f6/T--UESTC-China--plrecover.jpg" style="width:80% ;padding-top: 7%;"/></p>
<li>Dvorak, P., et al., Immobilized synthetic pathway for biodegradation of toxic recalcitrant pollutant 1,2,3-trichloropropane. Environ Sci Technol, 2014. 48(12): p. 6859-66.</li>
+
  <p class="mid">Figure 5. The main models of phytoremediation strategy.</p>
 
+
  <p>By understanding the physical and chemical properties of 1,2,3-TCP, we know that 1,2,3-TCP normally can not gather in living things and is easily to migrate due to its low adhesion. This makes it hard to deal with 1,2,3-TCP using the strategy of extraction and fixation. Moreover, phytostabilization and phytovolatilization requires frequent disposition and change of plants. This furthers requests to build another complete time and labor consuming system. phytoextraction won’t be taken into our consideration since 1,2,3-TCP can cause severe damage to human health by breathing. Therefore, we choose to use the strategy of phytodegradation to degrade 1,2,3-TCP into glycerol by creating super tobacco.</p>
<li>Pavlova, M., et al., Redesigning dehalogenase access tunnels as a strategy for degrading an anthropogenic substrate. Nat Chem Biol, 2009. 5(10): p. 727-33.</li>
+
 
+
<li>Wang, X., et al., Improvement of the thermostability and activity of halohydrin dehalogenase from Agrobacterium radiobacter AD1 by engineering C-terminal amino acids. J Biotechnol, 2015. 212: p. 92-8.</li>
+
 
+
<li>Buren, S., et al., Use of the foot-and-mouth disease virus 2A peptide co-expression system to study intracellular protein trafficking in Arabidopsis. PLoS One, 2012. 7(12): p. e51973.</li>
+
 
+
<li>Guo, A., J. Durner, and D.F. Klessig, Characterization of a tobacco epoxide hydrolase gene induced during the resistance response to TMV. Plant J, 1998. 15(5): p. 647-56.</li>
+
 
+
<li>Nitz, I., et al., Pyk10, a seedling and root specific gene and promoter from Arabidopsis thaliana. Plant Sci, 2001. 161(2): p. 337-346.</li>
+
 
+
<li>Werner, T., et al., Root-specific reduction of cytokinin causes enhanced root growth, drought tolerance, and leaf mineral enrichment in Arabidopsis and tobacco. Plant Cell, 2010. 22(12): p. 3905-20.</li>
+
 
+
<li>Nanasato, Y., et al., Biodegradation of gamma-hexachlorocyclohexane by transgenic hairy root cultures of Cucurbita moschata that accumulate recombinant bacterial LinA. Plant Cell Rep, 2016. 35(9): p. 1963-74.</li>
+
</ol>
+
 
+
 
</div>
 
</div>
 
+
<br /><br /><br /><br /><br />
+
  <div id="References">
</div>
+
    <h2>References</h2>
 +
    <ol>
 +
    <li>EPA. Technical Fact Sheet – 1,2,3-Trichloropropane (1,2,3-TCP), 2017.</li>
 +
    <li>Liu FS. The comprehensive utilization of 1,2,3-Trichloropropane. Speciality Petrochemicals, 1995;2:11-4.</li>
 +
    <li>Samin G, Janssen DB. Transformation and biodegradation of 1, 2, 3-trichloropropane (TCP). Environmental Science and Pollution Research, 2012. 1;19(8):3067-78.</li>
 +
 
 +
 +
    <li>Sasha Khokha . California Finally Begins Regulating Cancer-Causing Chemical Found in Drinking Water. KQED Science Menu, 2017.</li>
 +
  <li>Qian Yong. Research on Environment Behavior of 1,2,3-Trichloropropane in Groundwater of a Contaminated Site with Chlorinated Pollutants. China University of Geosciences(Beijing). 2016</li>
 +
 
 +
  <li>Tratnyek PG, Sarathy V, Fortuna JH. Fate and remediation of 1, 2, 3-trichloropropane. InInternational Conference on Remediation of Chlorinated and Recalcitrant Compounds, 6th, Monterey, CA 2008.</li>
 +
 +
  <li>Sarathy V, Salter AJ, Nurmi JT, O’Brien Johnson G, Johnson RL, Tratnyek PG. Degradation of 1, 2, 3-trichloropropane (TCP): hydrolysis, elimination, and reduction by iron and zinc. Environmental science & technology, 2009. 14;44(2):787-93.</li>
 +
 +
  <li>Sarathy V, Salter AJ, Nurmi JT, O’Brien Johnson G, Johnson RL, Tratnyek PG. Degradation of 1, 2, 3-trichloropropane (TCP): hydrolysis, elimination, and reduction by iron and zinc. Environmental science & technology, 2009. 14;44(2):787-93.</li>
 +
  <li>Kang JW. Removing environmental organic pollutants with bioremediation and phytoremediation. Biotechnology letters, 2014. 1;36(6):1129-39.</li>
 +
  <li>Cherian S, Oliveira MM. Transgenic plants in phytoremediation: recent advances and new possibilities. Environmental science & technology, 2005. 15;39(24):9377-90.</li>
 +
  <li>Kang JW. Removing environmental organic pollutants with bioremediation and phytoremediation. Biotechnology letters. 2014. 1;36(6):1129-39.</li>
 +
 +
    </ol>
 +
 +
 +
</div>
 +
 +
  </div>
 
</div>
 
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Latest revision as of 03:45, 2 November 2017

Team:UESTC-China/Introduction - 2017.igem.org

Overview

The progress of the chemical industry and agriculture has brought great convenience to our lives. But a large number of chemical pollutants which were toxic and difficult to degrade have been discharged to the rivers and soils, seriously polluting our environment. Xenobiotic organohalognse played a large part among all the chemical pollutants. After collecting a questionnaire for industrial and agricultural producers and doing a soil research in most areas of China, our team aimed at degrading an organic chloride, 1,2,3-Trichloropropane (1,2,3-TCP), which was less concerned by environmental protection department. In this summer, we decided to use synthetic biology to achieve plant degradation of 1,2,3-Trichloropropane by transferring three enzymes into tobacco which is environmentally friendly and the product, glycerol, is recyclable in tobacco.

The distributions of pollutant 1,2,3-TCP

1,2,3-TCP is an emerging organic pollutant which is usually been used as industrial solvents[1] and raw materials for producing 1,1,2,3-Tetrachloropropene or other chemicals substances[2]. In agriculture, it has been used as one of the ingredients of soil fumigants and it is also a harmful byproduct of other pesticides. The global production of 1,2,3-TCP reached about 50,000 tons annually[3]. Because 1,2,3-TCP is hard to degrade naturally in the environment, it would cause a huge damage to the groundwater and the soil if discharged casually.

One of the most serious pollution of 1,2,3-TCP was discovered in California. It has spread to all over the California since the 1940s when Dow Chemical and Shell started selling two soil fumigants (D-D and Telone) which include 1,2,3-TCP. Although 1,2,3-TCP was banned to use as soil fumigants in the 1990s, there was still a large amount of 1,2,3-TCP remained, to be a threat to the environment and people's lives seriously[4].

Figure 1. Water systems where significant levels of the 1,2,3-TCP have been detected in California

Dr. Yong Qian from University of Geosciences of China has explored the behavior of 1,2,3-TCP in the groundwater. He found 1,2,3-TCP still huge concentration (3890mg / L) underground in an abandoned factory that was only run from 1976 to 1979 in 2016 [5], which showed the great stability of 1,2,3-TCP in the groundwater and soil (Fig. 2).

Figure 2. The distribution of underground 1,2,3-TCP pollution in this factory [5].

Meanwhile, some studies showed that the adhesion coefficient of 1,2,3-TCP is very low[5]. It means that 1,2,3-TCP can be easily diffused into our daily use water. Its potential carcinogenicity and damage to kidney will threaten the health of human beings. In the recent past 10 years, more and more tests showing the existence of 1,2,3-TCP among worldwide drinking water is a sound proof (Fig. 3).

Figure 3. 1,2,3-TCP has been detected in hundreds of surface water and drinking water sources

Based on the above information, 1,2,3-TCP is absolutely a threatening pollutant. However, attentions are far more than enough to be paid on 1,2,3-TCP; and a lot of countries have not put 1,2,3-TCP as the test list of water quality. Since the current situation has been known to us, our team hopes to find a feasible way to stop 1,2,3-TCP by attracting the attentions of the whole society through doing advertisement and iGEM competition before it might cause great damage to the health of human beings.

Ways to solve the pollution of 1,2,3-TCP

Then the question is how to solve the problem of 1,2,3-TCP pollution? Traditional methods to deal with 1,2,3-TCP includes granular activated carbon (GAC)、hydrogen release compound (HRC)、reductive dechlorination by zero valent iron(ZVI) and son on[6]~[8]. However, some of these methods are either too inefficient or too expensive to be used within natural conditions (Fig. 4).

Figure 4. Treatability tests with 1,2,3-TCP-contaminated groundwater/soil

Since 1,2,3-TCP can hardly be dealt with traditional ways, Microbial remediation of 1,2,3-TCP is getting praised. Early studies have shown that 1,2,3-TCP can be converted into CO2、H2O and HCl by biological catalysis through Oxidation of metabolic mechanism with O2. So people are trying to find degradation method by using aerobe. But until now, all the tests to gather and filtrate aerobe to degrade 1,2,3-TCP have failed. However, scientists have found a few categories of bacteria that can degrade 1,2,3-TCP in absolute anaerobic environment. However, due to its strict requirement, the conversion efficiency is not high enough to get popular. In recent years, some studies have started to apply genetic engineering method to inject some enzymes in the seek of 1,2,3-TCP degradation. Some good results have been obtained but with some limitations. Firstly, these bacteria require very strict nutrition requirement and there only exits weak competition among them. Secondly, the diffusion of antibiotic resistance gene can be easily triggered. Finally, this method rely heavily on some specific induction condition[9]. We are not satisfied with these disadvantages and thus we hope to find a 1,2,3-TCP degradation method that is energy-efficient and sustainable. The burgeoning “Phytoremediation” has come into our attention.

Our super tobacco

As one of the new “Green remediation” strategy,phytoremediation shows enormous potential. Compared to microbial remediation, the most important advantage of phytoremediation is that phytoremediation comes with the potential to dispose pollutants by small amount of nutrition input. This is due to its unique feature of photosynthetic autotrophs system. By doing pollutant disposition, at the same time, plants can also help stabilize the soil, purify the water and clear air pollution[10]~[11]. There are four different ways of “Phytoremediation”: phytoextraction、phytostabilization、phytovolatilization and phytodegradation (Fig. 5).

Figure 5. The main models of phytoremediation strategy.

By understanding the physical and chemical properties of 1,2,3-TCP, we know that 1,2,3-TCP normally can not gather in living things and is easily to migrate due to its low adhesion. This makes it hard to deal with 1,2,3-TCP using the strategy of extraction and fixation. Moreover, phytostabilization and phytovolatilization requires frequent disposition and change of plants. This furthers requests to build another complete time and labor consuming system. phytoextraction won’t be taken into our consideration since 1,2,3-TCP can cause severe damage to human health by breathing. Therefore, we choose to use the strategy of phytodegradation to degrade 1,2,3-TCP into glycerol by creating super tobacco.

References

  1. EPA. Technical Fact Sheet – 1,2,3-Trichloropropane (1,2,3-TCP), 2017.
  2. Liu FS. The comprehensive utilization of 1,2,3-Trichloropropane. Speciality Petrochemicals, 1995;2:11-4.
  3. Samin G, Janssen DB. Transformation and biodegradation of 1, 2, 3-trichloropropane (TCP). Environmental Science and Pollution Research, 2012. 1;19(8):3067-78.
  4. Sasha Khokha . California Finally Begins Regulating Cancer-Causing Chemical Found in Drinking Water. KQED Science Menu, 2017.
  5. Qian Yong. Research on Environment Behavior of 1,2,3-Trichloropropane in Groundwater of a Contaminated Site with Chlorinated Pollutants. China University of Geosciences(Beijing). 2016
  6. Tratnyek PG, Sarathy V, Fortuna JH. Fate and remediation of 1, 2, 3-trichloropropane. InInternational Conference on Remediation of Chlorinated and Recalcitrant Compounds, 6th, Monterey, CA 2008.
  7. Sarathy V, Salter AJ, Nurmi JT, O’Brien Johnson G, Johnson RL, Tratnyek PG. Degradation of 1, 2, 3-trichloropropane (TCP): hydrolysis, elimination, and reduction by iron and zinc. Environmental science & technology, 2009. 14;44(2):787-93.
  8. Sarathy V, Salter AJ, Nurmi JT, O’Brien Johnson G, Johnson RL, Tratnyek PG. Degradation of 1, 2, 3-trichloropropane (TCP): hydrolysis, elimination, and reduction by iron and zinc. Environmental science & technology, 2009. 14;44(2):787-93.
  9. Kang JW. Removing environmental organic pollutants with bioremediation and phytoremediation. Biotechnology letters, 2014. 1;36(6):1129-39.
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