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measurement such as excessive noise signal in our sample.We therefore | measurement such as excessive noise signal in our sample.We therefore | ||
determined to test its function with a toxicity test. The product of PRK-RuBP | determined to test its function with a toxicity test. The product of PRK-RuBP | ||
− | cannot be metabolite by wild type E. coli. The accumulation of RuBP depletes | + | cannot be metabolite by wild type <i>E. coli</i>. The accumulation of RuBP depletes |
the sugar from the native pentose phosphate pathway. Lack of carbon source, the | the sugar from the native pentose phosphate pathway. Lack of carbon source, the | ||
growth of that strain may be repressed. We incubate the PRK expressing strain | growth of that strain may be repressed. We incubate the PRK expressing strain | ||
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<li>Check the growth and carbon fixation enhancement of CA enzyme</li> | <li>Check the growth and carbon fixation enhancement of CA enzyme</li> | ||
− | <li>Compare the carbon fixation rate of W3110 and BL21(DE3) E. coli strains</li> | + | <li>Compare the carbon fixation rate of W3110 and BL21(DE3) <i>E. coli</i> strains</li> |
<li>Compare different CO<sub>2</sub> incubation environment</li> | <li>Compare different CO<sub>2</sub> incubation environment</li> | ||
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<ol start="2"> | <ol start="2"> | ||
− | <li>The elemental formula of E. coli should be fixed or varies within a | + | <li>The elemental formula of <i>E. coli</i> should be fixed or varies within a |
small range. Although there may exist slightly different in different | small range. Although there may exist slightly different in different | ||
growth condition, we assume that such error can be ignore during the | growth condition, we assume that such error can be ignore during the | ||
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consumption over O.D. 600. O.D. 600 measurement can be viewed as the weight of | consumption over O.D. 600. O.D. 600 measurement can be viewed as the weight of | ||
carbon of the bacteria. The index shows the ratio of xylose consumption per | carbon of the bacteria. The index shows the ratio of xylose consumption per | ||
− | biomass. For wild type E. coli, it only consumes xylose (the sole carbon source | + | biomass. For wild type <i>E. coli</i>, it only consumes xylose (the sole carbon source |
− | provided in our medium) as its carbon source. Although some native E. coli | + | provided in our medium) as its carbon source. Although some native <i>E. coli</i> |
pathway | pathway | ||
may utilize CO<sub>2</sub> (such as lipid synthesis), the amount is too small to consider. | may utilize CO<sub>2</sub> (such as lipid synthesis), the amount is too small to consider. | ||
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<p class="pcontent"> | <p class="pcontent"> | ||
We then compare the XUI value between BL21(DE3) and W3110 constructed strain. | We then compare the XUI value between BL21(DE3) and W3110 constructed strain. | ||
− | When we design our IDT sequence, we link the CA directly to the promoter | + | When we design our IDT sequence, we link the CA directly to the promoter P<sub>LacI</sub>, |
so we could not transform CA construct into W3110 strain. We thus compare the | so we could not transform CA construct into W3110 strain. We thus compare the | ||
XUI of strains that only contains Rubisco and PRK (<a href="http://parts.igem.org/Part:BBa_K2762011" | XUI of strains that only contains Rubisco and PRK (<a href="http://parts.igem.org/Part:BBa_K2762011" | ||
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be lower with the expression of the constructed protein. The growth condition | be lower with the expression of the constructed protein. The growth condition | ||
of both constructed strains is similar for the first 12 hours. We then compare | of both constructed strains is similar for the first 12 hours. We then compare | ||
− | the difference of XUI between two E. coli strain. We found out that both strain | + | the difference of XUI between two <i>E. coli</i> strain. We found out that both strain |
shows similar trend: the XUI will be lower with the expression of the | shows similar trend: the XUI will be lower with the expression of the | ||
constructed protein. HoweverW3110 has a higher XUI compared with BL21(DE3) in | constructed protein. HoweverW3110 has a higher XUI compared with BL21(DE3) in | ||
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<div id="pt"> | <div id="pt"> | ||
<p class="pcontent"> | <p class="pcontent"> | ||
− | To find out how much and how efficient genetically engineered E. coli can fix | + | To find out how much and how efficient genetically engineered <i>E. coli</i> can fix |
carbon dioxide, we use the material balance concept to evaluate the | carbon dioxide, we use the material balance concept to evaluate the | ||
heterotrophic CO<sub>2</sub> fixation process. Consider a system composed of a single | heterotrophic CO<sub>2</sub> fixation process. Consider a system composed of a single | ||
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A system can be defined as an arbitrary portion of a process considered for | A system can be defined as an arbitrary portion of a process considered for | ||
− | analysis, in which in this case, is an engineered carbon capturing E. coli. | + | analysis, in which in this case, is an engineered carbon capturing <i>E. coli</i>. |
</p> | </p> | ||
</div> | </div> | ||
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<div id="pt"> | <div id="pt"> | ||
<p class="pcontent"> | <p class="pcontent"> | ||
− | The engineered E. coli BL21(DE3) are cultured in M9 medium with formula | + | The engineered </i>E. coli</i> BL21(DE3) are cultured in M9 medium with formula |
adjusted so that xylose is the sole carbon source. The aforementioned M9 Medium | adjusted so that xylose is the sole carbon source. The aforementioned M9 Medium | ||
contains | contains | ||
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ignored). By applying the law of conservation of mass, which states that mass | ignored). By applying the law of conservation of mass, which states that mass | ||
may neither be created nor destroyed, the material balance for carbon in an | may neither be created nor destroyed, the material balance for carbon in an | ||
− | engineered E. coli may simply be written as | + | engineered <i>E. coli</i> may simply be written as |
$${\{C_{CO_2}\ in\}\ +\ \{C_{xylose}\}\ -\ \{C_{CO_2}\ out\}\ -\ \{C_{waste}\}\ | $${\{C_{CO_2}\ in\}\ +\ \{C_{xylose}\}\ -\ \{C_{CO_2}\ out\}\ -\ \{C_{waste}\}\ | ||
=\ \{C_{biomass}\}...(1)}$$ | =\ \{C_{biomass}\}...(1)}$$ | ||
− | Considering the difficulties in measuring carbon in E. coli metabolic waste and | + | Considering the difficulties in measuring carbon in <i>E. coli</i> metabolic waste and |
that C<sub>waste</sub> would be positive, the equation reduces to | that C<sub>waste</sub> would be positive, the equation reduces to | ||
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<p class="pcontent"> | <p class="pcontent"> | ||
C<sub>biomass</sub> can be calculate by multiplying O.D. 600 to DCW and mass | C<sub>biomass</sub> can be calculate by multiplying O.D. 600 to DCW and mass | ||
− | percent of carbon in E. coli biomass. The O.D. 600 of engineered E. coli is | + | percent of carbon in <i>E. coli</i> biomass. The O.D. 600 of engineered <i>E. coli</i> is |
measured after a 12-hour cultivation and the result obtained is 0.45O.D. . Yin | measured after a 12-hour cultivation and the result obtained is 0.45O.D. . Yin | ||
− | Li et al. reported that dry cell weight (DCW) of E. coli is | + | Li et al. reported that dry cell weight (DCW) of <i>E. coli</i> is |
$${0.35g\over L ∙ 𝑂.𝐷. 600}$$ | $${0.35g\over L ∙ 𝑂.𝐷. 600}$$ | ||
− | , determined by experiment. E. coli biomass contains 48% of carbon by mass (or | + | , determined by experiment. <i>E. coli</i> biomass contains 48% of carbon by mass (or |
weight?). | weight?). | ||
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$${=\ 0.0066704\ g/L}$$ | $${=\ 0.0066704\ g/L}$$ | ||
− | Since the E. coli has been cultured for 12 hours, we can calculate the rate of | + | Since the <i>E. coli</i> has been cultured for 12 hours, we can calculate the rate of |
carbon fixation by | carbon fixation by | ||
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percent of carbon in biomass. | percent of carbon in biomass. | ||
− | $${Ratio\ of\ carbon\ in\ | + | $${Ratio\ of\ carbon\ in\ CO<sub>2</sub>\ fixed\ to\ carbon\ in\ biomass\ =\ |
{0.0066704\over 0.0756}}$$ | {0.0066704\over 0.0756}}$$ | ||
Revision as of 12:31, 14 October 2018
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