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| − | <h2>Transformation of <em>P. putida</em> KT2440</h2>
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| − | <p>
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| − | In order to avoid the virulence factors of
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| − | <em>Pseudomonas aeruginosa</em>, bacterial strains with
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| − | similar or shared metabolic pathways to the one
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| − | above were chosen as potential candidates. The
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| − | final candidates were <em>Pseudomonas putida</em> and
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| − | <em>Staphylococcus epidermidis</em>. Although
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| − | <em>S. epidermidis</em> doesn’t share the same exact
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| − | pathway as <em>P. aeruginosa</em>, it is a
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| − | naturally-occurring skin microbiome and only need
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| − | two additional enzymes, RhlA and RhlB, to produce
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| − | mono-rhamnolipids. Genes rhlA and rhlB necessary
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| − | for mono-rhamnolipid synthesis were extracted from
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| − | the <em>P. aeruginosa P14</em> bacterial strain. These
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| − | genes were cloned into the modified plasmid pNJ3.1
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| − | using standard cloning methods for transformation
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| − | into the desired bacterial strains (Figure 2). The
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| − | plasmid pC194 and a shuttle vector strain,
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| − | <em>S. aureus</em> RN4220 (details on <em>S. epidermidis</em>
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| − | transformation are discussed in the experiments
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| − | and result section) were used for <em>S. epidermidis</em>
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| − | transformations with the same basic design (Figure
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| − | 3). The conversion of mono-rhamnolipids to
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| − | di-rhamnolipids requires the additional gene rhlC,
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| − | which was also extracted from P14 strain and
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| − | cloned into the same pNJ3.1 vector (Figure 4).
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| − | </p>
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| − | <figure>
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| − | <img src="https://static.igem.org/mediawiki/parts/a/a4/RhlAB_circuit.png"
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| − | alt="RhlAB Plasmid Circuit" width="500">
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| − | </figure>
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| | ===Applications of BBa_K2062005=== | | ===Applications of BBa_K2062005=== |
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