IGEM:IMPERIAL/2009/Sun cream

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Sun Cream

Description

Development phases

Phase 1 - Compounds/threshold/detection

Tocochromanols are only synthesised in photosynthetic organisms; they are powerful antioxidants that protect the organism against reactive oxygen species (ROS). There has been successful reproduction of the biosynthetic pathway in E.coli through characterisation of the genes involved by knock-out and mutant studies. E.coli would be the chassis of choice.

Table from Ref 1:

Table 1. Strains and plasmids used in this work.

Strain or plasmid Relevant properties or genotype Source Strains
E. coli DH5F-, 80d, lacZ M15, endA1, recA1, hsdR17(rK-mK-), supE44, thi-1, gyrA96, relA1, (lacZYA-argF)U169 [a]
E. coli M15 [pREP4] F-, lacZ M15, thi-1, lac-, mtl-, recA+ , KmR [a]
Pseudomonas putida KT2440 type strain [b]
Synechocystis sp. PCC 6803 type strain [c]
pCAR16 -carotene biosynthesis gene cluster from Pantoea ananatis in pUC19, AmpR [27]
pJF119EH cloning vector, tac-promoter, IPTG inducible, AmpR [25]
pJF119 N cloning vector, RBS, tac-promoter, IPTG inducible, AmpR this study
pAW229 cloning vector, RBS, rha-promoter, L-rhamnose inducible, CmR [26]
pQE31-VTE1 tocopherol-cyclase (vte1) from Arabidopsis thaliana in pQE31, IPTG inducible, AmpR [18a]
pCAS2JF 1.2 kb EcoRI/BamHI PCR fragment hpd in pJF119EH this study
pCAS8 1.1 kb NdeI/BamHI PCR fragment ggh in pJF119 N this study
pCAS11 0.9 kb BglII/BamHI PCR fragment crtE in pCAS8 this study
pCAS12 1.0 kb BglII/BamHI PCR fragment hpt in pCAS11 this study
pCAS151.2 kb NdeI/BamHI PCR fragment hpd in pAW229 this study
pCAS18 1.2 kb BglII/BamHI PCR fragment hpd in pCAS11 this study
pCAS19 1.0 kb NdeI/BamHI PCR fragment hpt in pAW229 this study
pCAS23 1.0 kb NdeI/BamHI PCR fragment hpt in pJF119 N this study
pCAS24 0.9 kb NdeI/BamHI PCR fragment crtE in pCAS23 this study
pCAS27 1.1 kb NdeI/BamHI PCR fragment cyc-sy in pAW229 this study
pCAS29 1.2 kb BglII/BamHI PCR fragment hpd in pCAS18 this study
pCAS30 0.9 kb NdeI/BamHI PCR fragment crtE in pJF119 N this study
pCAS37 1.2 kb BglII/BamHI PCR fragment hpd in pCAS24 this study
pCAS47 1.3 kb BglII PCR fragment vte1 in pCAS29 this study


[a] Commercial strain. [b] Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ). [c] A kind gift from Prof. Forchhammer, Universität-Gießen (Germany).


Phase 2 - Encapsulation

Phase 3 - Killing strategy/gene deletion

Phase 4 - Storage

Phase 5 - Delivery

References

<html> <table> <tr> <td><A NAME="TBL1">&nbsp;</A></TD></TR> <tr> <td ALIGN="CENTER"><B>Table 1. </B><B>Strains and plasmids used in this work.</B></TD></TR> <tr VALIGN="BOTTOM"> <td> <hr size="2" noshade="1"></TD></TR> <tr> <td ALIGN="CENTER"> <table> <tr> <th ALIGN="left" VALIGN="TOP">

Strain or plasmid</TH> <th ALIGN="left" VALIGN="TOP">

Relevant properties or genotype</TH> <th ALIGN="left" VALIGN="TOP">

Source</TH></TR> <tr> <td COLSPAN="3"> <hr size="1"></TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<b>Strains</b></TD> <td ALIGN="left" VALIGN="TOP"></TD> <td ALIGN="left" VALIGN="TOP"></TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<I>E. coli</I> DH5<IMG SRC="/giflibrary/12/alpha.gif" BORDER="0"></TD> <td ALIGN="left" VALIGN="TOP">

F<SUP>-</SUP>, <I><IMG SRC="/giflibrary/12/phgr.gif" BORDER="0" ALIGN="ABSBOTTOM"></I>80d, <I>lacZ</I><IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">M15, <I>endA</I>1, <I>recA</I>1, <I>hsdR</I>17(r<SUB>K</SUB><SUP>-</SUP>m<SUB>K</SUB><SUP>-</SUP>), <I>supE</I>44, <I>thi</I>-1, <I>gyrA</I>96, <I>relA</I>1, <IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">(<I>lacZYA-argF</I>)U169</TD> <td ALIGN="left" VALIGN="TOP">

[a]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<I>E. coli</I> M15 [pREP4]</TD> <td ALIGN="left" VALIGN="TOP">

F<SUP>-</SUP>, <I>lacZ<IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">M15, thi</I>-1, <I>lac<SUP>-</SUP></I>, <I>mtl<SUP>-</SUP></I>, <I>recA<SUP>+</SUP></I> , Km<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

[a]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<I>Pseudomonas putida</I> KT2440</TD> <td ALIGN="left" VALIGN="TOP">

type strain</TD> <td ALIGN="left" VALIGN="TOP">

[b]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<I>Synechocystis</I> sp. PCC 6803</TD> <td ALIGN="left" VALIGN="TOP">

type strain</TD> <td ALIGN="left" VALIGN="TOP">

[c]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

<b>Plasmids</b></TD> <td ALIGN="left" VALIGN="TOP"></TD> <td ALIGN="left" VALIGN="TOP"></TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAR16</TD> <td ALIGN="left" VALIGN="TOP">

<IMG SRC="/giflibrary/12/beta.gif" BORDER="0" ALIGN="ABSBOTTOM">-carotene biosynthesis gene cluster from <I>Pantoea ananatis</I> in pUC19, Amp<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

[<A HREF="#BIB27">27</A>]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pJF119EH</TD> <td ALIGN="left" VALIGN="TOP">

cloning vector, tac-promoter, IPTG inducible, Amp<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

[<A HREF="#BIB25">25</A>]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pJF119<IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">N</TD> <td ALIGN="left" VALIGN="TOP">

cloning vector, RBS, tac-promoter, IPTG inducible, Amp<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pAW229</TD> <td ALIGN="left" VALIGN="TOP">

cloning vector, RBS, <I>rha</I>-promoter, <FONT SIZE="-1">L</FONT>-rhamnose inducible, Cm<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

[<A HREF="#BIB26">26</A>]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pQE31-VTE1</TD> <td ALIGN="left" VALIGN="TOP">

tocopherol-cyclase (vte1) from <I>Arabidopsis thaliana</I> in pQE31, IPTG inducible, Amp<SUP>R</SUP></TD> <td ALIGN="left" VALIGN="TOP">

[<A HREF="#BIB18A">18a</A>]</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS2JF</TD> <td ALIGN="left" VALIGN="TOP">

1.2 kb EcoRI/BamHI PCR fragment <I>hpd</I> in pJF119EH</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS8</TD> <td ALIGN="left" VALIGN="TOP">

1.1 kb NdeI/BamHI PCR fragment <I>ggh</I> in pJF119<IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">N</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS11</TD> <td ALIGN="left" VALIGN="TOP">

0.9 kb BglII/BamHI PCR fragment <I>crtE</I> in pCAS8</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS12</TD> <td ALIGN="left" VALIGN="TOP">

1.0 kb BglII/BamHI PCR fragment <I>hpt</I> in pCAS11</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS15</TD> <td ALIGN="left" VALIGN="TOP">

1.2 kb NdeI/BamHI PCR fragment <I>hpd</I> in pAW229</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS18</TD> <td ALIGN="left" VALIGN="TOP">

1.2 kb BglII/BamHI PCR fragment <I>hpd</I> in pCAS11</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS19</TD> <td ALIGN="left" VALIGN="TOP">

1.0 kb NdeI/BamHI PCR fragment <I>hpt</I> in pAW229</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS23</TD> <td ALIGN="left" VALIGN="TOP">

1.0 kb NdeI/BamHI PCR fragment <I>hpt</I> in pJF119<IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">N</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS24</TD> <td ALIGN="left" VALIGN="TOP">

0.9 kb NdeI/BamHI PCR fragment <I>crtE</I> in pCAS23</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS27</TD> <td ALIGN="left" VALIGN="TOP">

1.1 kb NdeI/BamHI PCR fragment <I>cyc-sy</I> in pAW229</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS29</TD> <td ALIGN="left" VALIGN="TOP">

1.2 kb BglII/BamHI PCR fragment <I>hpd</I> in pCAS18</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS30</TD> <td ALIGN="left" VALIGN="TOP">

0.9 kb NdeI/BamHI PCR fragment <I>crtE</I> in pJF119<IMG SRC="/giflibrary/12/Delta.gif" BORDER="0">N</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS37</TD> <td ALIGN="left" VALIGN="TOP">

1.2 kb BglII/BamHI PCR fragment <I>hpd</I> in pCAS24</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR> <tr> <td ALIGN="left" VALIGN="TOP">

pCAS47</TD> <td ALIGN="left" VALIGN="TOP">

1.3 kb BglII PCR fragment <I>vte1</I> in pCAS29</TD> <td ALIGN="left" VALIGN="TOP">

this study</TD></TR></TABLE></TD></TR> <tr VALIGN="BOTTOM"> <td> <hr size="2" noshade="1"></TD></TR></TABLE></CENTER><CENTER> <table width="80%"> <tr> <td><FONT SIZE="-1">[a] Commercial strain. [b] Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ). [c] A kind gift from Prof. Forchhammer, Universit&auml;t-Gie&szlig;en (Germany). <br></FONT></TD></TR></TABLE> </html>

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