Difference between revisions of "Part:BBa K4263006"

 
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glyceraldehyde-3-phosphate-dehydrogenase) promoter (PGAP), one of the strongest constitutive promoter in P. pastoris
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===Usage and Biology===
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<span class='h3bb'>Sequence and Features</span>
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    <h2 style="font-weight: bold;">P<sub>GAP</sub></h2>
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    <h2>Introduction</h2>
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    <p>Glyceraldehyde-3-phosphate-dehydrogenase (GAP) promoter is one of the strongest constitutive promoter in <em>Pichia pastoris</em><sup>[1]</sup>.</p>
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    <h2>Characterization</h2>
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===Functional Parameters===
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    <p>In order to test the function of P<em><sub>GAP</sub></em>, we construct "P<em><sub>GAP</sub>-EGFP-</em>terminator" (Figure 1). If P<em><sub>GAP</sub></em> is functional, we can test the fluorescence intensity of EGFP in supernatant samples obtained from the culture of recombinant <em>P.pastoris</em> GS115 strain.</p>
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    <img src="https://static.igem.wiki/teams/4263/wiki/parts/image/gape-min.jpg" alt="">
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    <h4>Figure 1 Gene circuit of P<em><sub>GAP</sub>-EGFP-</em>terminator</h4>
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    <p>Our results matched the general expected trend (Figure 2). After fermentation experiment in BMMY medium containing 1% methanol. The fluorescence intensity of the samples of recombinant <em>P.pastoris</em> GS115 containing the <em>EGFP</em> gene was essentially unchanged over time. At the same time, we measured the growth curve of the strains.</p>
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    <img src="https://static.igem.wiki/teams/4263/wiki/parts/image/pgap-min.jpg" alt="">
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    <h4>Figure 2 Fluorescence intensity and OD600 absorbance of samples obtained at different time points from the culture of corresponding recombinant <em>P.pastoris</em> GS115 containing <em>EGFP</em> gene.</h4>
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    <h2>Reference</h2>
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    <p>[1] Waterham, H.R., Digan, M.E., Koutz, P.J., Lair, S.V., Cregg, J.M., 1997. Isolation of the <em>Pichia pastoris</em> glyceraldehyde-3-phosphate dehydrogenase gene and regulation and use of its promoter. Gene 186 (1), 37-44.</p>
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Revision as of 15:41, 26 September 2022

<!DOCTYPE html> K4263006

PGAP

Introduction

Glyceraldehyde-3-phosphate-dehydrogenase (GAP) promoter is one of the strongest constitutive promoter in Pichia pastoris[1].

Characterization

In order to test the function of PGAP, we construct "PGAP-EGFP-terminator" (Figure 1). If PGAP is functional, we can test the fluorescence intensity of EGFP in supernatant samples obtained from the culture of recombinant P.pastoris GS115 strain.

Figure 1 Gene circuit of PGAP-EGFP-terminator

Our results matched the general expected trend (Figure 2). After fermentation experiment in BMMY medium containing 1% methanol. The fluorescence intensity of the samples of recombinant P.pastoris GS115 containing the EGFP gene was essentially unchanged over time. At the same time, we measured the growth curve of the strains.

Figure 2 Fluorescence intensity and OD600 absorbance of samples obtained at different time points from the culture of corresponding recombinant P.pastoris GS115 containing EGFP gene.

Reference

[1] Waterham, H.R., Digan, M.E., Koutz, P.J., Lair, S.V., Cregg, J.M., 1997. Isolation of the Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase gene and regulation and use of its promoter. Gene 186 (1), 37-44.