http://2007.igem.org/wiki/index.php?title=Special:Contributions/Sergio_Peisajovich&feed=atom&limit=50&target=Sergio_Peisajovich&year=&month=
2007.igem.org - User contributions [en]
2024-03-28T12:04:04Z
From 2007.igem.org
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http://2007.igem.org/wiki/index.php/UCSF
UCSF
2007-10-26T21:36:08Z
<p>Sergio Peisajovich: /* A Guided Tour through the Project: JUDGES, THIS IS FOR YOU! */</p>
<hr />
<div><div id="mainpage"><br />
[[Image:logo6.png]]<br />
</div><br />
<br />
{| cell spacing=“1”<br />
|width="300px" bgcolor="gray" |<br />
<!-- PEOPLE HEADER with edit link--><br />
<h3><html><img src=http://openwetware.org/images/3/39/Icon_groups.png alt=People><br />
</html><font color=black>People<br />
<hr/><br />
<!-- END PEOPLE HEADER with edit link--><br />
===Students===<br />
*[[Michael Chen]] (UC Berkeley)<br />
*[[Eric Chou]] (Lincoln High)<br />
*[[Jimmy Huang]] (Lincoln High)<br />
*[[Lauren Jann]] (Lincoln High)<br />
*[[Eric Meltzer]] (Palo Alto High)<br />
*[[Alexander Ng]] (Lincoln High)<br />
*[[Robert Ovadia]] (Lincoln High)<br />
<br />
===Instructors===<br />
*[[Noah Helman]]<br />
*[[Andrew Horwitz]]<br />
*[[Sergio Peisajovich]]<br />
*[[David Pincus]]<br />
*[[Ryan Ritterson]]<br />
*[[Nili Sommovilla]]<br />
*[[Reid Williams]]<br />
*[[Angela Won]]<br />
<br />
===Faculty Advisors===<br />
*[http://www.ucsf.edu/limlab/ Wendell Lim]<br />
*[[Hana El-Samad]]<br />
*[http://kortemmelab.ucsf.edu/ Tanja Kortemme]<br />
*[http://www.voigtlab.ucsf.edu/ Chris Voigt] <br />
</font><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
|width="280px" bgcolor="gainsboro" |<br />
<!-- COMMUNITY HEADER with edit link--><br />
<html><h3><br />
<img src="http://openwetware.org/images/b/b9/Icon_board.png" alt=Resources><br />
</html>The Project</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
===About Us===<br />
*[[/Our Story/]]<br />
===A Guided Tour through the Project: JUDGES, THIS IS FOR YOU!===<br />
*[[/Project Description/]]<br />
*[[/Main Introduction/]]<br />
*'''Project One: Engineering Protein Complexes'''<br />
**[[/Intro/]]<br />
**[[/Results/]]<br />
**[[/Conclusions/]]<br />
*'''Project Two: The Synthetic Organelle'''<br />
**[[/Organelle Intro/| Intro]]<br />
**[[/Results_P2/| Results]]<br />
**[[/Conclusions_P2/| Status]]<br />
*[[/Conclusions/| Overall Conclusions]]<br />
<br />
===Behind the Scenes===<br />
*'''Methods and Materials'''<br />
**'''Project One'''<br />
***[[/Cloning Strategy1/| Cloning Strategy]]<br />
***[[/YeastStrains1/| Yeast Strains]]<br />
***[[/Flow Cytometry1/| Flow Cytometry]]<br />
**'''Project Two'''<br />
***[[/Cloning Strategy2/| Cloning Strategy]]<br />
***[[/YeastStrains2/|Yeast Strains]]<br />
***[[/Flow Cytometry2/|Flow Cytometry]]<br />
***[[/Microscopy/|Microscopy]]<br />
*[[IGEM:UCSF/2007/Plasmids | Plasmids & Sequences]]<br />
<br />
<br />
<br />
<br />
<br />
<br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
|width="300px" bgcolor="gray" | <br />
<!-- NEWS HEADER with edit link--><br />
<font color=black><h3><html><br />
<img src="http://openwetware.org/images/e/e2/Icon_info.png" alt="News" border=”0”><br />
</html>Information</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
<br />
===Planning & Development===<br />
<br />
*[[/Weekly Meetings/]]<br />
*[[/Past Presentations/]]<br />
*[[/Papers of Interest/]]<br />
*[[Logo Galore...]]<br />
<br />
===Media===<br />
*[[IGEM:UCSF/2007/Photos | Photos]]<br />
*[[/Videos/]]<br />
*[http://169.230.117.36/today/cache/feature/200708022.html in ucsf news!]<br />
<br />
===iGEM links===<br />
Tools<br />
*[http://www.pubmed.org PubMed]<br />
*[http://www.yeastgenome.org Yeast DB]<br />
*[http://openwetware.org/wiki/Protocols OpenWetWare Protocols]<br />
For Visitors<br />
*[http://www.ucsf.edu UCSF Homepage]<br />
*[http://partsregistry.org Registry of Parts]<br />
*[https://2007.igem.org/wiki/index.cgi iGEM 2007]<br />
*Who's been checking us out?<br />
<html> <a href="http://www3.clustrmaps.com/counter/maps.php?url=http://openwetware.org/wiki/IGEM:UCSF/2007"><img src="http://www3.clustrmaps.com/stats/maps-no_clusters/openwetware.org-wiki-IGEM-UCSF-2007-thumb.jpg" /><br />
</a> </html></font><br />
<br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
__NOTOC__</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF
UCSF
2007-10-26T21:35:18Z
<p>Sergio Peisajovich: /* A Guided Tour through the Project: JUDGES, THIS IS FOR YOU! */</p>
<hr />
<div><div id="mainpage"><br />
[[Image:logo6.png]]<br />
</div><br />
<br />
{| cell spacing=“1”<br />
|width="300px" bgcolor="gray" |<br />
<!-- PEOPLE HEADER with edit link--><br />
<h3><html><img src=http://openwetware.org/images/3/39/Icon_groups.png alt=People><br />
</html><font color=black>People<br />
<hr/><br />
<!-- END PEOPLE HEADER with edit link--><br />
===Students===<br />
*[[Michael Chen]] (UC Berkeley)<br />
*[[Eric Chou]] (Lincoln High)<br />
*[[Jimmy Huang]] (Lincoln High)<br />
*[[Lauren Jann]] (Lincoln High)<br />
*[[Eric Meltzer]] (Palo Alto High)<br />
*[[Alexander Ng]] (Lincoln High)<br />
*[[Robert Ovadia]] (Lincoln High)<br />
<br />
===Instructors===<br />
*[[Noah Helman]]<br />
*[[Andrew Horwitz]]<br />
*[[Sergio Peisajovich]]<br />
*[[David Pincus]]<br />
*[[Ryan Ritterson]]<br />
*[[Nili Sommovilla]]<br />
*[[Reid Williams]]<br />
*[[Angela Won]]<br />
<br />
===Faculty Advisors===<br />
*[http://www.ucsf.edu/limlab/ Wendell Lim]<br />
*[[Hana El-Samad]]<br />
*[http://kortemmelab.ucsf.edu/ Tanja Kortemme]<br />
*[http://www.voigtlab.ucsf.edu/ Chris Voigt] <br />
</font><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
|width="280px" bgcolor="gainsboro" |<br />
<!-- COMMUNITY HEADER with edit link--><br />
<html><h3><br />
<img src="http://openwetware.org/images/b/b9/Icon_board.png" alt=Resources><br />
</html>The Project</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
===About Us===<br />
*[[/Our Story/]]<br />
===A Guided Tour through the Project: JUDGES, THIS IS FOR YOU!===<br />
*[[/Project Description/]]<br />
*[[/Main Introduction/]]<br />
*'''Project One: Engineering Protein Complexes'''<br />
**[[/Intro/]]<br />
**[[/Results/]]<br />
**[[/Conclusions/]]<br />
*'''Project Two: The Synthetic Organelle'''<br />
**[[/Organelle Intro/| Intro]]<br />
**[[/Results_P2/| Results]]<br />
**[[/Conclusions_P2/| Status]]<br />
**[[/Conclusions_P2/| Overall Conclusions]]<br />
<br />
===Behind the Scenes===<br />
*'''Methods and Materials'''<br />
**'''Project One'''<br />
***[[/Cloning Strategy1/| Cloning Strategy]]<br />
***[[/YeastStrains1/| Yeast Strains]]<br />
***[[/Flow Cytometry1/| Flow Cytometry]]<br />
**'''Project Two'''<br />
***[[/Cloning Strategy2/| Cloning Strategy]]<br />
***[[/YeastStrains2/|Yeast Strains]]<br />
***[[/Flow Cytometry2/|Flow Cytometry]]<br />
***[[/Microscopy/|Microscopy]]<br />
*[[IGEM:UCSF/2007/Plasmids | Plasmids & Sequences]]<br />
<br />
<br />
<br />
<br />
<br />
<br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
|width="300px" bgcolor="gray" | <br />
<!-- NEWS HEADER with edit link--><br />
<font color=black><h3><html><br />
<img src="http://openwetware.org/images/e/e2/Icon_info.png" alt="News" border=”0”><br />
</html>Information</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
<br />
===Planning & Development===<br />
<br />
*[[/Weekly Meetings/]]<br />
*[[/Past Presentations/]]<br />
*[[/Papers of Interest/]]<br />
*[[Logo Galore...]]<br />
<br />
===Media===<br />
*[[IGEM:UCSF/2007/Photos | Photos]]<br />
*[[/Videos/]]<br />
*[http://169.230.117.36/today/cache/feature/200708022.html in ucsf news!]<br />
<br />
===iGEM links===<br />
Tools<br />
*[http://www.pubmed.org PubMed]<br />
*[http://www.yeastgenome.org Yeast DB]<br />
*[http://openwetware.org/wiki/Protocols OpenWetWare Protocols]<br />
For Visitors<br />
*[http://www.ucsf.edu UCSF Homepage]<br />
*[http://partsregistry.org Registry of Parts]<br />
*[https://2007.igem.org/wiki/index.cgi iGEM 2007]<br />
*Who's been checking us out?<br />
<html> <a href="http://www3.clustrmaps.com/counter/maps.php?url=http://openwetware.org/wiki/IGEM:UCSF/2007"><img src="http://www3.clustrmaps.com/stats/maps-no_clusters/openwetware.org-wiki-IGEM-UCSF-2007-thumb.jpg" /><br />
</a> </html></font><br />
<br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
__NOTOC__</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/Jamboree/Schedule/Friday/Practice_talks
Jamboree/Schedule/Friday/Practice talks
2007-10-26T20:52:17Z
<p>Sergio Peisajovich: /* Friday 2 Nov : Practice Talks sign-up sheet */</p>
<hr />
<div>== Friday 2 Nov : Practice Talks sign-up sheet ==<br />
<br />
<br />
Use this sign-up sheet to sign up for a slot on Friday night to practice your talk. Note that there will NOT be any A/V (audio/visual) support on staff. All classrooms will be unlocked and you should use them and leave them as you found them. <br />
<br />
<br />
(Pizza and refreshments will be available on a first-come first-serve basis)<br />
<br />
<br />
<html><br />
<link rel="stylesheet" href="https://2007.igem.org/wiki/index.php?title=User:Macowell/schedule.css&action=raw&ctype=text/css"><br />
<table class="calendar"><h2 class="date"><a name="Friday Practice">Friday, Nov 2</a></h2><br />
<thead><br />
<tr><br />
<th width="15%">Time</th><br />
<th>room 123</th><br />
<th>room 124</th><br />
<th>room 141</th><br />
<th>room 144</th><br />
<th>room 155</th><br />
</tr><br />
</thead><br />
<tbody><br />
<tr class="even"><br />
<th>6:00p - 6:30p</th><br />
<td>A1 - Glasgow</td><br />
<td>B1 - Cambridge (UK)</td><br />
<td>C1</td><br />
<td>D1</td><br />
<td>E1</td><br />
</tr><br />
<tr class="odd"><br />
<th>6:30p - 7:00p</th><br />
<td>A2 </td><br />
<td>B2</td><br />
<td>C2</td><br />
<td>D2</td><br />
<td>E2</td><br />
</tr><br />
<tr class="even"><br />
<th>7:00p - 7:30p</th><br />
<td>A3</td><br />
<td>B3</td><br />
<td>C3</td><br />
<td>D3</td><br />
<td>E3</td><br />
</tr><br />
<tr class="even"><br />
<th>7:30p - 8:00p</th><br />
<td>A3</td><br />
<td>B4</td><br />
<td>C4</td><br />
<td>D4</td><br />
<td>E4</td><br />
</tr><br />
<tr class="odd"><br />
<th>8:00p - 8:30p</th><br />
<td>McGill</td><br />
<td>B5</td><br />
<td>C5</td><br />
<td>D5</td><br />
<td>E5 - Taipei</td><br />
</tr><br />
<tr class="even"><br />
<th>8:30p - 9:00p</th><br />
<td>A6</td><br />
<td>B6</td><br />
<td>C6</td><br />
<td>D6</td><br />
<td>E6</td><br />
</tr><br />
<tr class="odd"><br />
<th>9:00p - 9:30p</th><br />
<td>A7</td><br />
<td>UCSF</td><br />
<td>C7</td><br />
<td>D7</td><br />
<td>E7</td><br />
</tr><br />
<tr class="even"><br />
<th>9:30p - 10:00p</th><br />
<td>A8</td><br />
<td>Davidson / Mo West</td><br />
<td>C8</td><br />
<td>D8</td><br />
<td>E8</td><br />
</tr><br />
</tbody><br />
</table><br />
</html></div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/IGEM:UCSF/2007/Plasmids
IGEM:UCSF/2007/Plasmids
2007-10-26T20:12:49Z
<p>Sergio Peisajovich: </p>
<hr />
<div><u>'''PROJECT 1</u><br />
<br />
<u>'''Expected sequences</u><br />
<br />
''Below you have the sequences of the different parts in the right order, as well as the adaptor sequences between parts. You will need to arrange them accordingly to your particular constructs, so that you will have 1-2-3-4-5-6-7-8.''<br />
<br />
<br />
<br />
'''1-Promoter sequence (variable, depends on particular acceptor)'''<br />
<br />
<br />
pFig1<br />
<br />
TTGAATGATCAACCAAACGCCGATATAGTCACTATTGGTGATAAAATTAAAATGC<br />
AGCTATTGAGAACCCATCCTAACAACATTAAATTGTCTATTAATGGAATTGGTAA<br />
AGATGCCCCAACTTTCCAAGAATCTGCTTTGATTGCCGATAAGTTATTGAGTGTC<br />
ATGAAGGCCGGCACTTACCCAAAGATTTCCATTTTCTACAATGACCCAGTGTCTT<br />
CCCTATCTTTTGAACCATCTGAAAAACCGATCTTTAACGCCAAGACCATTGAACA<br />
ATCCCCATCATTCGGCAAATTTGAGATCGACACGGACGCAAACGTTCCAAGAGAT<br />
TTGTTTGAATATACTTTGGCTAACCAAATGTTGACAGCAATGGCTCAAGGTTATG<br />
CTGCTGAAATTTCCGCCAGAAGAAACGCTATGGATAACGCTTCCAAGAATGCCGG<br />
TGATATGATCAATCGTTACTCTATCTTGTACAACAGAACAAGACAAGCTGTCATT<br />
ACTAATGAACTGGTTGATATTATTACTGGTGCTTCCTCTTTGGGATGATAAAAAA<br />
ATCACCCTGCATTGCCTCTTTATTTGACGTTGTTTTTGTAGAACATGAAACGAAT<br />
TTTGACTTGATGAGACGAAGTATATATCCAAAGAATACCTTAAATAGAAAAGGAA<br />
AGATAATAAATACTAAACACTACTATATATTCAGGTAAAATACAAAAATTATAAC<br />
ATTTTTAAAACTTTTTTTTTTTGAAAGTCCTTCTCGCTTTAGGATTTTTTCCCAT<br />
TAAGATTATGATGGTTTCATGTATGTGTCAGTTAAAAAAAATATGGCTAAGTAGC<br />
AATGAAACGAACCAAGAAAATGAAAACAATATATAGTGCTGTTGAAATAACAAAG<br />
ACATTGGTATATATTTGTAAAATGTCTGTTAAATGTTTTTTATCTCAGGTTCTTG<br />
CTTGTCTTTGGTAGAAGAAATTATAGTAAACAAACAAACAAACAAACAAAAAAAA<br />
AAAAAAAAAA<br />
<br />
pAdh<br />
<br />
TTAAAACAAGAAGAGGGTTGACTACATCACGATGAGGGGGATCGAAGAAATGATG<br />
GTAAATGAAATAGGAAATCAAGGAGCATGAAGGCAAAAGACAAATATAAGGGTCG<br />
AACGAAAAATAAAGTGAAAAGTGTTGATATGATGTATTTGGCTTTGCGGCGCCGA<br />
AAAAACGAGTTTACGCAATTGCACAATCATGCTGACTCTGTGGCGGACCCGCGCT<br />
CTTGCCGGCCCGGCGATAACGCTGGGCGTGAGGCTGTGCCCGGCGGAGTTTTTTG<br />
CGCCTGCATTTTCCAAGGTTTACCCTGCGCTAAGGGGCGAGATTGGAGAAGCAAT<br />
AAGAATGCCGGTTGGGGTTGCGATGATGACGACCACGACAACTGGTGTCATTATT<br />
TAAGTTGCCGAAAGAACCTGAGTGCATTTGCAACATGAGTATACTAGAAGAATGA<br />
GCCAAGACTTGCGAGACGCGAGTTTGCCGGTGGTGCGAACAATAGAGCGACCATG<br />
ACCTTGAAGGTGAGACGCGCATAACCGCTAGAGTACTTTGAAGAGGAAACAGCAA<br />
TAGGGTTGCTACCAGTATAAATAGACAGGTACATACAACACTGGAAATGGTTGTC<br />
TGTTTGAGTACGCTTTCAATTCATTTGGGTGTGCACTTTATTATGTTACAATATG<br />
GAAGGGAACTTTACACTTCTCCTATGCACATATATTAATTAAAGTCCAATGCTAG<br />
TAGAGAAGGGGGGTAACACCCCTCCGCGCTCTTTTCCGATTTTTTTCTAAACCGT<br />
GGAATATTTCGGATATCCTTTTGTTGTTTCCGGGTGTACAATATGGACTTCCTCT<br />
TTTCTGGCAACCAAACCCATACATCGGGATTCCTATAATACCTTCGTTGGTCTCC<br />
CTAACATGTAGGTGGCGGAGGGGAGATATACAATAGAACAGATACCAGACAAGAC<br />
ATAATGGGCTAAACAAGACTACACCAATTACACTGCCTCATTGATGGTGGTACAT<br />
AACGAACTAATACTGTAGCCCTAGACTTGATAGCCATCATCATATCGAAGTTTCA<br />
CTACCCTTTTTCCATTTGCCATCTATTGAAGTAATAATAGGCGCATGCAACTTCT<br />
TTTCTTTTTTTTTCTTTTCTCTCTCCCCCGTTGTTGTCTCACCATATCCGCAATG<br />
ACAAAAAAATGATGGAAGACACTAAAGGAAAAAATTAACGACAAAGACAGCACCA<br />
ACAGATGTCGTTGTTCCAGAGCTGATGAGGGGTATCTCGAAGCACACGAAACTTT<br />
TTCCTTCCTTCATTCACGCACACTACTCTCTAATGAGCAACGGTATACGGCCTTC<br />
CTTCCAGTTACTTGAATTTGAAATAAAAAAAAGTTTGCTGTCTTGCTATCAAGTA<br />
TAAATAGACCTGCAATTATTAATCTTTTGTTTCCTCGTCATTGTTCTCGTTCCCT<br />
TTCTTCCTTGTTTCTTTTTCTGCACAATATTTCAAGCTATACCAAGCATACAATC<br />
AACTATCTCATATACA<br />
<br />
<br />
pCyc<br />
<br />
CAGATCCGCCAGGCGTGTATATATAGCGTGGATGGCCAGGCAACTTTAGTGCTGA<br />
CACATACAGGCATATATATATGTGTGCGACGACACATGATCATATGGCATGCATG<br />
TGCTCTGTATGTATATAAAACTCTTGTTTTCTTCTTTTCTCTAAATATTCTTTCC<br />
TTATACATTAGGACCTTTGCAGCATAAATTACTATACTTCTATAGACACACAAAC<br />
ACAAATACACACACTAAATTAATA<br />
<br />
<br />
pSte5<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCT<br />
CTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTT<br />
CCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAG<br />
AAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAG<br />
TAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCAC<br />
ATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTA<br />
ACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGG<br />
CGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAA<br />
AGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAA<br />
AGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTA<br />
GCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAA<br />
<br />
<br />
'''2-ATG-XhoI-“A” and 2 adaptor bases:'''<br />
ATGCTCGAGGGAGCT<br />
<br />
<br />
'''3-mCherry:'''<br />
<br />
GTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGGAGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACG<br />
AGTTCGAGATCGAGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCAAGGGTGGCCCCCTGCC<br />
CTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCAAGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAG<br />
CTGTCCTTCCCCGAGGGCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGGACTCCTCCCTGC<br />
AGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCCCCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTG<br />
GGAGGCCTCCTCCGAGCGGATGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACGGCGGCCAC<br />
TACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGCCCGGCGCCTACAACGTCAACATCAAGTTGGACATCA<br />
CCTCCCACAACGAGGACTACACCATCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGTACAAG <br />
<br />
<br />
<br />
'''4-Adaptor bases, followed by “B” and another adaptor base:'''<br />
GGTAGTTCCCTA <br />
<br />
<br />
<br />
'''5-Effectors'''<br />
<br />
For YopH:<br />
<br />
AACTTATCATTAAGCGATCTTCATCGTCAGGTATCTCGATTGGTGCAGCAAGAGAGCGGTGATTGTACCGGGAAATTAAGAGGTAACGT<br />
TGCTGCCAATAAAGAAACTACCTTTCAAGGTTTGACCATAGCCAGTGGAGCCAGAGAGTCAGAAAAAGTATTTGCTCAAACTGTACTAA<br />
GCCACGTAGCAAATGTTGTTCTAACTCAAGAAGATACCGCTAAGCTATTGCAAAGTACGGTAAAGCATAATTTGAATAATTATGACTTA<br />
AGAAGTGTCGGCAATGGTAATAGTGTACTTGTCAGTTTACGTAGTGACCAAATGACACTACAAGACGCCAAAGTGCTGTTGGAGGCCGC<br />
ATTGCGACAAGAGTCGGGAGCGAGGGGGCATGTATCATCTCATTCACATTCAGCCCTTCACGCACCGGGAACCCCGGTGCGTGAAGGAC<br />
TGCGTTCACATCTAGACCCCAGAACTCCACCGTTGCCACCGCGTGAACGACCACACACTTCTGGCCATCACGGGGCTGGCGAAGCCAGA<br />
GCCACCGCACCAAGCACTGTTTCTCCTTATGGCCCAGAAGCGCGCGCAGAACTCAGCAGCCGCCTCACCACATTGCGCAATACGCTGGC<br />
GCCAGCAACGAATGATCCGCGTTACTTACAAGCCTGCGGCGGTGAAAAGCTAAACCGATTTAGAGATATTCAATGCTGTCGGCAAACCG<br />
CAGTACGCGCCGATCTTAATGCCAATTACATCCAGGTCGGTAACACTCGTACCATAGCGTGCCAGTATCCGCTACAATCTCAACTTGAA<br />
AGCCATTTCCGTATGCTGGCAGAAAACCGAACGCCAGTGTTGGCTGTTTTAGCGTCCAGTTCTGAGATAGCCAATCAAAGATTCGGTAT<br />
GCCAGATTATTTCCGCCAGAGTGGTACCTATGGCAGTATCACTGTAGAGTCTAAAATGACTCAGCAAGTTGGTCTCGGTGACGGGATTA<br />
TGGCAGATATGTATACTTTAACGATTCGTGAAGCGGGTCAAAAAACAATCTCTGTTCCTGTGGTTCATGTTGGCAATTGGCCCGATCAG<br />
ACCGCAGTCAGCTCTGAAGTTACCAAGGCACTCGCTTCACTGGTAGATCAAACAGCAGAAACAAAACGCAATATGTATGAAAGCAAAGG<br />
AAGTTCAGCGGTAGGAGATGACTCCAAATTACGGCCGGTAATACATTGCCGTGCGGGTGTTGGCCGTACTGCGCAACTGATTGGCGCAA<br />
TGTGCATGAATGATAGTCGTAATAGTCAGTTAAGCGTAGAAGATATGGTCAGCCAAATGCGAGTACAAAGAAATGGTATTATGGTACAA<br />
AAAGATGAGCAACTTGATGTTCTGATTAAGTTGGCTGAAGGACAAGGGCGACCATTATTAAATAGC<br />
<br />
For YopJ:<br />
<br />
ATCGGACCAATATCACAAATAAATATCTCCGGTGGCTTATCAGAAAAAGAGACCAGTTCTTTAATCAGTAATGAAGAGCTTAAAAATAT<br />
CATAACACAGTTGGAAACTGATATATCGGATGGATCCTGGTTCCATAAAAATTATTCACGTATGGATGTAGAAGTCATGCCCGCATTGG<br />
TAATCCAGGCGAACAATAAATATCCGGAAATGAATCTTAATCTTGTTACATCTCCATTGGACCTTTCAATAGAAATAAAAAACGTCATA<br />
GAAAATGGAGTTAGATCTTCCCGCTTCATAATTAACATGGGGGAAGGTGGAATACATTTCAGTGTAATTGATTACAAACATATAAATGG<br />
GAAAACATCTCTGATATTGTTTGAACCAGCAAACTTTAACAGTATGGGGCCAGCGATGCTGGCAATAAGGACAAAAACGGCTATTGAAC<br />
GTTATCAATTACCTGATTGCCATTTCTCCATGGTGGAAATGGATATTCAGCGAAGCTCATCTGAATGTGGTATTTTTAGTTTGGCACTG<br />
GCAAAAAAACTTTACATCGAGAGAGATAGCCTGTTGAAAATACATGAAGATAATATAAAAGGTATATTAAGTGATGGTGAAAATCCTTT<br />
ACCCCACGATAAGTTGGACCCGTATCTCCCGGTAACTTTTTACAAACATACTCAAGGTAAAAAACGTCTTAATGAATATTTAAATACTA<br />
ACCCGCAGGGAGTTGGTACTGTTGTTAACAAAAAAAATGAAACCATCGTTAATAGATTTGATAACAATAAATCCATTGTAGATGGAAAG<br />
GATTATCAGTTTCGGTACATAAAAAGAGAATAGCTGAATATAAAACACTTCTCAAAGTA<br />
<br />
For OspF:<br />
<br />
CCAATTAAAAAGCCTTGTTTGAAATTGAATTTAGATTCTTTGAACGTTGTTAGATCAGAAATTCCACAAATGTTATCTGCTAATGAAAGAT<br />
TGAAAAATAATTTTAACATTTTATATAATCAAATAAGACAGTACCCAGCTTATTATTTTAAAGTTGCTTCTAATGTTCCTACTTACTCTGA<br />
TATATGTCAATTTTTCTCAGTTATGTATCAAGGTTTTCAGATAGTTAATCATTCTGGTGATGTTTTTATTCATGCTTGTAGAGAAAATCCA<br />
CAATCTAAAGGTGATTTTGTTGGTGACAAGTTCCATATTTCTATAGCTAGAGAACAAGTTCCATTGGCATTTCAAATTTTATCTGGTTTGTT<br />
ATTTTCTGAGGACTCTCCTATTGACAAATGGAAGATAACTGATATGAATAGAGTTTCTCAACAGTCAAGAGTTGGTATTGGTGCTCAATTTAC<br />
TTTGTATGTTAAATCTGATCAAGAATGTTCTCAGTATTCAGCTTTATTGTTGCATAAAATTAGACAATTCATAATGTGCTTGGAATCTAATTT<br />
GTTAAGATCTAAGATTGCTCCAGGTGAATATCCTGCATCTGATGTTAGACCAGAGGATTGGAAGTACGTTTCATATAGAAATGAATTGAGATC<br />
TGACAGAGATGGTTCTGAAAGACAAGAGCAGATGTTAAGAGAAGAACCTTTTTATAGATTGATGATTGAG<br />
<br />
<br />
<br />
'''6-Adaptor bases, followed by “C” followed by another adaptor:'''<br />
GGTAGCGAT<br />
<br />
<br />
<br />
'''7-Zippers:'''<br />
For RR:<br />
GGTTCTGGTTCTGGTTCTGGTTCTCTGGAAATCCGTGCGGCGTTCCTGGAAAAAGAAAACACCGCGCTGCGTACCCGTGCGGCGGAACTGCG<br />
TAAACGTGTTGGTCGTTGCCGTAACATCGTTTCTAAATACGAAACCCGTTACGGTCCGCTG<br />
For EE:<br />
<br />
GGTTCCGGTAGCGGTTCCGGTTCTATCACTATTAGGGCTGCTTTTTTGGAAAAAGAAAATACTGCTTTGAGAACTGAAATTGCTGAATTGG<br />
AAAAAGAAGTTGGTAGATGTGAAAATATTGTTTCTAAATATGAAACTAGATATGGTCCATTG<br />
<br />
<br />
<br />
'''8-Final adaptors, followed by “D”, followed by BamHI, 3 STOPS in three different frames, then NotI, the Adh terminator and SacI:'''<br />
<br />
<br />
TAATGCGGGATCCTGAGTAAATAAGCGGCCGCGCGAATTTC<br />
TTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTAT<br />
ACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGT<br />
AACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT<br />
ATTGACCACACCGAGCTC<br />
<br />
<br />
<u>'''PROJECT 2</u><br />
<br />
<br />
(1) PspOMI//pSte2-Ste2//XhoI//GFP//XmaI//RR//STOP//BamHI//AdhTerminator<br />
<br />
GGGCCCATCCAATATCACCTGACCTTCATCTTTATGTTTTTCCACTGGTGTAATGTACTCTGTTGCATCATCAATAAGCAACTGGGACGCCAA<br />
TAATTCATACCAACAATCTTTTCTTAATGAGTTGTTTACAAACCCCAGAGGCCCTAAACCTATGTACCTCAAAGCTCGGACATCTTTTTTCTC<br />
TAATAACTTTGATATAACTTTTCCTTTTTGTTCTTCTGCCTTTCCACTGTCACAACGAGAAAGTGATTCCAGATATGCGTTATAACCTCCCCT<br />
TGTCAACGCGTCTTTGTCGTGGCTACTCTGATTAGTATGAAATAATGACCTTAATGGCATCGTATTTTGTTAATTGGCTTGTTCTATGTCATT<br />
TGCTAAATTTTCTCTTTATTTCTTTACTTTGCCAATTCAAGATAATCTTCCCTTCCCAGAGAGAAAAAAGGAAAAATTTAGCTATGAAACCTC<br />
AATAAGCTTTTTAATACACCAAAGATTCAAGATAAGAGCATAGAACGAACTGTAGAATAGTCCGGATATGTTATCCAATGCCTGCCAAAATGC<br />
ATTGTCACACGCTGTAGTGCTCGAATAGGTGTTGCAATCCGTCAATATACGTCTTGCTCTGTGGGTAAATGTCTCGTGCATTAAGACAGGCTA<br />
GTATAAACGAGAAGAAGTATCCTGCTTTGCAATGAAACAATAGTATCCGCTAAGAATTTAAGCAGGCCAACGTCCATACTGCTTAGGACCTGT<br />
GCCTGGCAAGTCGCAGATTGAAGTTTTTTCAACCATGTAAATTTCCTAATTGGGTAAGTACATGATGAAACACATATGAAGAAAAAAGCTTTC<br />
CTACATATTCAAGATTTTTTTCTGTGGGTGGAATACTATTTAAGGAGTGCTATTAGTATCTTATTTGACTTCAAAGCAATACGATACCTTTTC<br />
TTTTCACCTGCTCTGGCTATAATTATAATTGGTTACTTAAAAATGCACCGTTAAGAACCATATCCAAGAATCAAAAATGTCTGATGCGGCTCC<br />
TTCATTGAGCAATCTATTTTATGATCCAACGTATAATCCTGGTCAAAGCACCATTAACTACACTTCCATATATGGGAATGGATCTACCATCAC<br />
TTTCGATGAGTTGCAAGGTTTAGTTAACAGTACTGTTACTCAGGCCATTATGTTTGGTGTCAGATGTGGTGCAGCTGCTTTGACTTTGATTGT<br />
CATGTGGATGACATCGAGAAGCAGAAAAACGCCGATTTTCATTATCAACCAAGTTTCATTGTTTTTAATCATTTTGCATTCTGCACTCTATTT<br />
TAAATATTTACTGTCTAATTACTCTTCAGTGACTTACGCTCTCACCGGATTTCCTCAGTTCATCAGTAGAGGTGACGTTCATGTTTATGGTGC<br />
TACAAATATAATTCAAGTCCTTCTTGTGGCTTCTATTGAGACTTCACTGGTGTTTCAGATAAAAGTTATTTTCACAGGCGACAACTTCAAAAG<br />
GATAGGTTTGATGCTGACGTCGATATCTTTCACTTTAGGGATTGCTACAGTTACCATGTATTTTGTAAGCGCTGTTAAAGGTATGATTGTGAC<br />
TTATAATGATGTTAGTGCCACCCAAGATAAATACTTCAATGCATCCACAATTTTACTTGCATCCTCAATAAACTTTATGTCATTTGTCCTGGT<br />
AGTTAAATTGATTTTAGCTATTAGATCAAGAAGATTCCTTGGTCTCAAGCAGTTCGATAGTTTCCATATTTTACTCATAATGTCATGTCAATC<br />
TTTGTTGGTTCCATCGATAATATTCATCCTCGCATACAGTTTGGAACCAAACCAGGGAACAGATGTCTTGACTACTGTTGCAACATTACTTGC<br />
TGTATTGTCTTTACCATTATCATCAATGTGGGCCACGGCTGCTAATAATGCATCCAAAACAAACACAATTACTTCAGACTTTACAACATCCAC<br />
AGATAGGTTTTATCCAGGCACGCTGTCTAGCTTTCAAACTGATAGTATCAACAACGATGCTAAAAGCAGTCTCAGAAGTAGATTATATGACCT<br />
ATATCCTAGAAGGAAGGAAACAACATCGGATAAACATTCGGAAAGAACTTTTGTTTCTGAGACTGCAGATGATATAGAGAAAAATCAGTTTTA<br />
TCAGTTGCCCACACCTACGAGTTCAAAAAATACTAGGATAGGACCGTTTGCTGATGCAAGTTACAAAGAGGGAGAAGTTGAACCCGTCGACAT<br />
GTACACTCCCGATACGGCAGCTGATGAGGAAGCCAGAAAGTTCTGGACTGAAGATAATAATAATTTACTCGAGTCTAAAGGTGAAGAATTATT<br />
CACTGGTGTTGTCCCAATTTTGGTTGAATTAGATGGTGATGTTAATGGTCACAAATTTTCTGTCTCCGGTGAAGGTGAAGGTGATGCTACTTA<br />
CGGTAAATTGACCTTAAAATTTATTTGTACTACTGGTAAATTGCCAGTTCCATGGCCAACCTTAGTCACTACTTTAACTTATGGTGTTCAATG<br />
TTTTTCTAGATACCCAGATCATATGAAACAACATGACTTTTTCAAGTCTGCCATGCCAGAAGGTTATGTTCAAGAAAGAACTATTTTTTTCAA<br />
AGATGACGGTAACTACAAGACCAGAGCTGAAGTCAAGTTTGAAGGTGATACCTTAGTTAATAGAATCGAATTAAAAGGTATTGATTTTAAAGA<br />
AGATGGTAACATTTTAGGTCACAAATTGGAATACAACTATAACTCTCACAATGTTTACATCATGGCTGACAAACAAAAGAATGGTATCAAAGT<br />
TAACTTCAAAATTAGACACAACATTGAAGATGGTTCTGTTCAATTAGCTGACCATTATCAACAAAATACTCCAATTGGTGATGGTCCAGTCTT<br />
GTTACCAGACAACCATTACTTATCCACTCAATCTaggTTATCCAAAGATCCAAACGAAAAGAGAGACCACATGGTCTTGTTAGAATTTGTTAC<br />
TGCTGCTGGTATTACCCATGGTATGGATGAATTGTACAAACCCGGGggatctggatcgggatctGATCCAGATTTGGAAATTAGAGCTGCTTT<br />
TTTGAGACAAAGAAATACTGCTTTGAGAACTGAAGTTGCTGAATTGGAACAAGAAGTTCAAAGATTGGAAAATGAAGTTTCTCAATATGAAAC<br />
TAGATATGGTCCATTGGGTGGTGGTAAATAAGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAA<br />
GTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGT<br />
ATAGCATGAGGTCGCTCTTATTGACCACACC<br />
<br />
<br />
(2) PspOMI//pGal1//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
TGTTATTGTGATAGTATCTCAGCGGGATTTGTGTAGATTCACCTACAGCCTTTAAACCAGCAAAGTGAATTACCGAATCAATTTTATATTCTT<br />
TGAAAACCTTTTCCAGACCTTTTCGGTCACACAAATCAACCTCATAGAAGGGAATGTGATGCTTGGTCAAGACCTCTAACCTGGCTACAGAAT<br />
CATAAGTTGAATTCGACAGGTTATCAGCAACAACACAGTCATATCCATTCTCAATTAGCTCTACCACAGTGTGTGAACCAATGTATCCAGCAC<br />
CACCTGTAACCAAAACAATTTTAGAAGTACTTTCACTTTGTAACTGAGCTGTCATTTATATTGAATTTTCAAAAATTCTTACTTTTTTTTTGG<br />
ATGGACGCAAAGAAGTTTAATAATCATATTACATGGCATTACCACCATATACATATCCATATCTAATCTTACTTATATGTTGTGGAAATGTAA<br />
AGAGCCCCATTATCTTAGCCTAAAAAAACCTTCTCTTTGGAACTTTCAGTAATACGCTTAACTGCTCATTGCTATATTGAAGTACGGATTAGA<br />
AGCCGCCGAGCGGGCGACAGCCCTCCGACGGAAGACTCTCCTCCGTGCGTCCTCGTCTTCACCGGTCGCGTTCCTGAAACGCAGATGTGCCTC<br />
GCGCCGCACTGCTCCGAACAATAAAGATTCTACAATACTAGCTTTTATGGTTATGAAGAGGAAAAATTGGCAGTAACCTGGCCCCACAAACCT<br />
TCAAATTAACGAATCAAATTAACAACCATAGGATGATAATGCGATTAGTTTTTTAGCCTTATTTCTGGGGTAATTAATCAGCGAAGCGATGAT<br />
TTTTGATCTATTAACAGATATATAAATGGAAAAGCTGCATAACCACTTTAACTAATACTTTCAACATTTTCAGTTTGTATTACTTCTTATTCA<br />
AATGTCATAAAAGTATCAACAAAAAATTGTTAATATACCTCTATACTTTAACGTCAAGGAGAAAAAACTATAATGCTCGAGGGAGCTgcttct<br />
gcatcaacttctaaatataattcacactccttggagaatgagtctattaagaggacgtctcgagatggagtcaatcgagatctcactgaggct<br />
gttcctcgacttccaggagaaacactaatcactgacaaagaagttatttacatatgtcctttcaatggccccattaagggaagagtttacatc<br />
acaaattatcgtctttatttaagaagtttggaaacggattcttctctaatacttgatgttcctctgggtgtgatctcgagaattgaaaaaatg<br />
ggaggcgcgacaagtagaggagaaaattcctatggtctagatattacttgtaaagacatgagaaacctgaggttcgctttgaaacaggaaggc<br />
cacagcagaagagatatgtttgagatcctcacgagatacgcgtttcccctggctcacagtctgccattatttgcatttttaaatgaagaaaag<br />
tttaacgtggatggatggacagtttacaatccagtggaagaatacaggaggcagggcttgcccaatcaccattggagaataacttttattaat<br />
aagtgctatgagctctgtgacacttaccctgctcttttggtggttccgtatcgtgcctcagatgatgacctccggagagttgcaacttttagg<br />
tcccgaaatcgaattccagtgctgtcatggattcatccagaaaataagacggtcattgtgcgttgcagtcagcctcttgtcggtatgagtggg<br />
aaacgaaataaagatgatgagaaatatctcgatgttatcagggagactaataaacaaatttctaaactcaccatttatgatgcaagacccagc<br />
gtaaatgcagtggccaacaaggcaacaggaggaggatatgaaagtgatgatgcatatcataacgccgaacttttcttcttagacattcataat<br />
attcatgttatgcgggaatctttaaaaaaagtgaaggacattgtttatcctaatgtagaagaatctcattggttgtccagtttggagtctact<br />
cattggttagaacatatcaagctcgttttgacaggagccattcaagtagcagacaaagtttcttcagggaagagttcagtgcttgtgcattgc<br />
agtgacggatgggacaggactgctcagctgacatccttggccatgctgatgttggatagcttctataggagcattgaagggttcgaaatactg<br />
gtacaaaaagaatggataagttttggacataaatttgcatctcgaataggtcatggtgataaaaaccacaccgatgctgaccgttctcctatt<br />
tttctccagtttattgattgtgtgtggcaaatgtcaaaacagttccctacagcttttgaattcaatgaacaatttttgattataattttggat<br />
catctgtatagttgccgatttggtactttcttattcaactgtgaatctgctcgagaaagacagaaggttacagaaaggactgtttctttatgg<br />
tcactgataaacagtaataaagaaaaattcaaaaaccccttctatactaaagaaatcaatcgagttttatatccagttgccagtatgcgtcac<br />
ttggaactctgggtgaattactacattagatggaaccccaggatcaagcaacaacagccgaatccagtggagcagcgttacatggagctctta<br />
gccttacgcgacgaatacataaagcggcttgaggaactgcagctcgccaactctgccaagctttctgatcccccaacttcaccttccagtcct<br />
tcgcaaatgatgccccatgtgcaaactcacttcGGTAGTTCCCTAggttccggtagcggttccggttctatcactattagggctgcttttttg<br />
gaaaaagaaaatactgctttgagaactgaaattgctgaattggaaaaagaagttggtagatgtgaaaatattgtttctaaatatgaaactaga<br />
tatggtccattgTAATCGCGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAAT<br />
TTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGT<br />
CGCTCTTATTGACCACACC<br />
<br />
(3) pGal1//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but WMTM1 sequence<br />
<br />
(4) pSte5//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but Ste5 promoter sequence:<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCTCTGATAACACGTTGTTTGAACATCGACAAGATGAAAATC<br />
TAGAAGTATCAAGTTTCCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAGAAGAAGCGTTAAATATTGGATCT<br />
TTCCGCAGTTCTACTCTGATACATTTTTGAAGTAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCACATGAATT<br />
AATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTAACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTT<br />
TGGGTGCGGCGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAAAGGCGATAGTAGCTAAAGAAAATACCGAGA<br />
ATTTCCTCGAAAAGTTGACGACAAAAGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTAGCATCCGCGCTAAA<br />
AAAGGAAGATACAGGATACAGCGGAAACAACTTTTAAATG<br />
<br />
(5) pSte5//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above.<br />
<br />
(6) pCyc//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but Cyc promoter sequence:<br />
<br />
ctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgaca<br />
catgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttg<br />
cagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatg<br />
<br />
(7) pCyc//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above.<br />
<br />
(8) pCyc//ATG//XhoI//GFP//XmaI//(SerGlySerGlySer)ING2 PH domain//STOP//BamHI<br />
<br />
As above, but //(SerGlySerGlySer)ING2 PH domain sequence as follows:<br />
<br />
TCTGGATCAGGTTCAGCTTCTCCAGTTGAATTTGCTATTGATCCTAACGAACCTACATACTGCTTATGTAATCAAGTTTCTTATGGTGAAATGAT<br />
TGGTTGCGATAACGAGCAGTGTCCAATAGAATGGTTTCATTTCTCTTGCGTTTCTTTAACATATAAACCTAAGGGAAAATGGTACTGTCCTAAAT<br />
GTAGAGGTGATAATGAGAAAACTATGGATAAGTCTACTGAAAAGACAAAAAAAGATAGAAGATCAAGA</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/IGEM:UCSF/2007/Plasmids
IGEM:UCSF/2007/Plasmids
2007-10-26T20:08:21Z
<p>Sergio Peisajovich: </p>
<hr />
<div><u>'''PROJECT 1</u><br />
<br />
<u>'''Expected sequences</u><br />
<br />
''Below you have the sequences of the different parts in the right order, as well as the adaptor sequences between parts. You will need to arrange them accordingly to your particular constructs, so that you will have 1-2-3-4-5-6-7-8.''<br />
<br />
<br />
<br />
'''1-Promoter sequence (variable, depends on particular acceptor)'''<br />
<br />
<br />
pFig1<br />
<br />
TTGAATGATCAACCAAACGCCGATATAGTCACTATTGGTGATAAAATTAAAATGC<br />
AGCTATTGAGAACCCATCCTAACAACATTAAATTGTCTATTAATGGAATTGGTAA<br />
AGATGCCCCAACTTTCCAAGAATCTGCTTTGATTGCCGATAAGTTATTGAGTGTC<br />
ATGAAGGCCGGCACTTACCCAAAGATTTCCATTTTCTACAATGACCCAGTGTCTT<br />
CCCTATCTTTTGAACCATCTGAAAAACCGATCTTTAACGCCAAGACCATTGAACA<br />
ATCCCCATCATTCGGCAAATTTGAGATCGACACGGACGCAAACGTTCCAAGAGAT<br />
TTGTTTGAATATACTTTGGCTAACCAAATGTTGACAGCAATGGCTCAAGGTTATG<br />
CTGCTGAAATTTCCGCCAGAAGAAACGCTATGGATAACGCTTCCAAGAATGCCGG<br />
TGATATGATCAATCGTTACTCTATCTTGTACAACAGAACAAGACAAGCTGTCATT<br />
ACTAATGAACTGGTTGATATTATTACTGGTGCTTCCTCTTTGGGATGATAAAAAA<br />
ATCACCCTGCATTGCCTCTTTATTTGACGTTGTTTTTGTAGAACATGAAACGAAT<br />
TTTGACTTGATGAGACGAAGTATATATCCAAAGAATACCTTAAATAGAAAAGGAA<br />
AGATAATAAATACTAAACACTACTATATATTCAGGTAAAATACAAAAATTATAAC<br />
ATTTTTAAAACTTTTTTTTTTTGAAAGTCCTTCTCGCTTTAGGATTTTTTCCCAT<br />
TAAGATTATGATGGTTTCATGTATGTGTCAGTTAAAAAAAATATGGCTAAGTAGC<br />
AATGAAACGAACCAAGAAAATGAAAACAATATATAGTGCTGTTGAAATAACAAAG<br />
ACATTGGTATATATTTGTAAAATGTCTGTTAAATGTTTTTTATCTCAGGTTCTTG<br />
CTTGTCTTTGGTAGAAGAAATTATAGTAAACAAACAAACAAACAAACAAAAAAAA<br />
AAAAAAAAAA<br />
<br />
pAdh<br />
<br />
TTAAAACAAGAAGAGGGTTGACTACATCACGATGAGGGGGATCGAAGAAATGATG<br />
GTAAATGAAATAGGAAATCAAGGAGCATGAAGGCAAAAGACAAATATAAGGGTCG<br />
AACGAAAAATAAAGTGAAAAGTGTTGATATGATGTATTTGGCTTTGCGGCGCCGA<br />
AAAAACGAGTTTACGCAATTGCACAATCATGCTGACTCTGTGGCGGACCCGCGCT<br />
CTTGCCGGCCCGGCGATAACGCTGGGCGTGAGGCTGTGCCCGGCGGAGTTTTTTG<br />
CGCCTGCATTTTCCAAGGTTTACCCTGCGCTAAGGGGCGAGATTGGAGAAGCAAT<br />
AAGAATGCCGGTTGGGGTTGCGATGATGACGACCACGACAACTGGTGTCATTATT<br />
TAAGTTGCCGAAAGAACCTGAGTGCATTTGCAACATGAGTATACTAGAAGAATGA<br />
GCCAAGACTTGCGAGACGCGAGTTTGCCGGTGGTGCGAACAATAGAGCGACCATG<br />
ACCTTGAAGGTGAGACGCGCATAACCGCTAGAGTACTTTGAAGAGGAAACAGCAA<br />
TAGGGTTGCTACCAGTATAAATAGACAGGTACATACAACACTGGAAATGGTTGTC<br />
TGTTTGAGTACGCTTTCAATTCATTTGGGTGTGCACTTTATTATGTTACAATATG<br />
GAAGGGAACTTTACACTTCTCCTATGCACATATATTAATTAAAGTCCAATGCTAG<br />
TAGAGAAGGGGGGTAACACCCCTCCGCGCTCTTTTCCGATTTTTTTCTAAACCGT<br />
GGAATATTTCGGATATCCTTTTGTTGTTTCCGGGTGTACAATATGGACTTCCTCT<br />
TTTCTGGCAACCAAACCCATACATCGGGATTCCTATAATACCTTCGTTGGTCTCC<br />
CTAACATGTAGGTGGCGGAGGGGAGATATACAATAGAACAGATACCAGACAAGAC<br />
ATAATGGGCTAAACAAGACTACACCAATTACACTGCCTCATTGATGGTGGTACAT<br />
AACGAACTAATACTGTAGCCCTAGACTTGATAGCCATCATCATATCGAAGTTTCA<br />
CTACCCTTTTTCCATTTGCCATCTATTGAAGTAATAATAGGCGCATGCAACTTCT<br />
TTTCTTTTTTTTTCTTTTCTCTCTCCCCCGTTGTTGTCTCACCATATCCGCAATG<br />
ACAAAAAAATGATGGAAGACACTAAAGGAAAAAATTAACGACAAAGACAGCACCA<br />
ACAGATGTCGTTGTTCCAGAGCTGATGAGGGGTATCTCGAAGCACACGAAACTTT<br />
TTCCTTCCTTCATTCACGCACACTACTCTCTAATGAGCAACGGTATACGGCCTTC<br />
CTTCCAGTTACTTGAATTTGAAATAAAAAAAAGTTTGCTGTCTTGCTATCAAGTA<br />
TAAATAGACCTGCAATTATTAATCTTTTGTTTCCTCGTCATTGTTCTCGTTCCCT<br />
TTCTTCCTTGTTTCTTTTTCTGCACAATATTTCAAGCTATACCAAGCATACAATC<br />
AACTATCTCATATACA<br />
<br />
<br />
pCyc<br />
<br />
CAGATCCGCCAGGCGTGTATATATAGCGTGGATGGCCAGGCAACTTTAGTGCTGA<br />
CACATACAGGCATATATATATGTGTGCGACGACACATGATCATATGGCATGCATG<br />
TGCTCTGTATGTATATAAAACTCTTGTTTTCTTCTTTTCTCTAAATATTCTTTCC<br />
TTATACATTAGGACCTTTGCAGCATAAATTACTATACTTCTATAGACACACAAAC<br />
ACAAATACACACACTAAATTAATA<br />
<br />
<br />
pSte5<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCT<br />
CTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTT<br />
CCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAG<br />
AAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAG<br />
TAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCAC<br />
ATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTA<br />
ACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGG<br />
CGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAA<br />
AGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAA<br />
AGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTA<br />
GCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAA<br />
<br />
<br />
'''2-ATG-XhoI-“A” and 2 adaptor bases:'''<br />
ATGCTCGAGGGAGCT<br />
<br />
<br />
'''3-mCherry:'''<br />
<br />
GTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGGAGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACG<br />
AGTTCGAGATCGAGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCAAGGGTGGCCCCCTGCC<br />
CTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCAAGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAG<br />
CTGTCCTTCCCCGAGGGCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGGACTCCTCCCTGC<br />
AGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCCCCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTG<br />
GGAGGCCTCCTCCGAGCGGATGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACGGCGGCCAC<br />
TACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGCCCGGCGCCTACAACGTCAACATCAAGTTGGACATCA<br />
CCTCCCACAACGAGGACTACACCATCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGTACAAG <br />
<br />
<br />
<br />
'''4-Adaptor bases, followed by “B” and another adaptor base:'''<br />
GGTAGTTCCCTA <br />
<br />
<br />
<br />
'''5-Effectors'''<br />
<br />
For YopH:<br />
<br />
AACTTATCATTAAGCGATCTTCATCGTCAGGTATCTCGATTGGTGCAGCAAGAGAGCGGTGATTGTACCGGGAAATTAAGAGGTAACGT<br />
TGCTGCCAATAAAGAAACTACCTTTCAAGGTTTGACCATAGCCAGTGGAGCCAGAGAGTCAGAAAAAGTATTTGCTCAAACTGTACTAA<br />
GCCACGTAGCAAATGTTGTTCTAACTCAAGAAGATACCGCTAAGCTATTGCAAAGTACGGTAAAGCATAATTTGAATAATTATGACTTA<br />
AGAAGTGTCGGCAATGGTAATAGTGTACTTGTCAGTTTACGTAGTGACCAAATGACACTACAAGACGCCAAAGTGCTGTTGGAGGCCGC<br />
ATTGCGACAAGAGTCGGGAGCGAGGGGGCATGTATCATCTCATTCACATTCAGCCCTTCACGCACCGGGAACCCCGGTGCGTGAAGGAC<br />
TGCGTTCACATCTAGACCCCAGAACTCCACCGTTGCCACCGCGTGAACGACCACACACTTCTGGCCATCACGGGGCTGGCGAAGCCAGA<br />
GCCACCGCACCAAGCACTGTTTCTCCTTATGGCCCAGAAGCGCGCGCAGAACTCAGCAGCCGCCTCACCACATTGCGCAATACGCTGGC<br />
GCCAGCAACGAATGATCCGCGTTACTTACAAGCCTGCGGCGGTGAAAAGCTAAACCGATTTAGAGATATTCAATGCTGTCGGCAAACCG<br />
CAGTACGCGCCGATCTTAATGCCAATTACATCCAGGTCGGTAACACTCGTACCATAGCGTGCCAGTATCCGCTACAATCTCAACTTGAA<br />
AGCCATTTCCGTATGCTGGCAGAAAACCGAACGCCAGTGTTGGCTGTTTTAGCGTCCAGTTCTGAGATAGCCAATCAAAGATTCGGTAT<br />
GCCAGATTATTTCCGCCAGAGTGGTACCTATGGCAGTATCACTGTAGAGTCTAAAATGACTCAGCAAGTTGGTCTCGGTGACGGGATTA<br />
TGGCAGATATGTATACTTTAACGATTCGTGAAGCGGGTCAAAAAACAATCTCTGTTCCTGTGGTTCATGTTGGCAATTGGCCCGATCAG<br />
ACCGCAGTCAGCTCTGAAGTTACCAAGGCACTCGCTTCACTGGTAGATCAAACAGCAGAAACAAAACGCAATATGTATGAAAGCAAAGG<br />
AAGTTCAGCGGTAGGAGATGACTCCAAATTACGGCCGGTAATACATTGCCGTGCGGGTGTTGGCCGTACTGCGCAACTGATTGGCGCAA<br />
TGTGCATGAATGATAGTCGTAATAGTCAGTTAAGCGTAGAAGATATGGTCAGCCAAATGCGAGTACAAAGAAATGGTATTATGGTACAA<br />
AAAGATGAGCAACTTGATGTTCTGATTAAGTTGGCTGAAGGACAAGGGCGACCATTATTAAATAGC<br />
<br />
For YopJ:<br />
<br />
ATCGGACCAATATCACAAATAAATATCTCCGGTGGCTTATCAGAAAAAGAGACCAGTTCTTTAATCAGTAATGAAGAGCTTAAAAATAT<br />
CATAACACAGTTGGAAACTGATATATCGGATGGATCCTGGTTCCATAAAAATTATTCACGTATGGATGTAGAAGTCATGCCCGCATTGG<br />
TAATCCAGGCGAACAATAAATATCCGGAAATGAATCTTAATCTTGTTACATCTCCATTGGACCTTTCAATAGAAATAAAAAACGTCATA<br />
GAAAATGGAGTTAGATCTTCCCGCTTCATAATTAACATGGGGGAAGGTGGAATACATTTCAGTGTAATTGATTACAAACATATAAATGG<br />
GAAAACATCTCTGATATTGTTTGAACCAGCAAACTTTAACAGTATGGGGCCAGCGATGCTGGCAATAAGGACAAAAACGGCTATTGAAC<br />
GTTATCAATTACCTGATTGCCATTTCTCCATGGTGGAAATGGATATTCAGCGAAGCTCATCTGAATGTGGTATTTTTAGTTTGGCACTG<br />
GCAAAAAAACTTTACATCGAGAGAGATAGCCTGTTGAAAATACATGAAGATAATATAAAAGGTATATTAAGTGATGGTGAAAATCCTTT<br />
ACCCCACGATAAGTTGGACCCGTATCTCCCGGTAACTTTTTACAAACATACTCAAGGTAAAAAACGTCTTAATGAATATTTAAATACTA<br />
ACCCGCAGGGAGTTGGTACTGTTGTTAACAAAAAAAATGAAACCATCGTTAATAGATTTGATAACAATAAATCCATTGTAGATGGAAAG<br />
GATTATCAGTTTCGGTACATAAAAAGAGAATAGCTGAATATAAAACACTTCTCAAAGTA<br />
<br />
For OspF:<br />
<br />
CCAATTAAAAAGCCTTGTTTGAAATTGAATTTAGATTCTTTGAACGTTGTTAGATCAGAAATTCCACAAATGTTATCTGCTAATGAAAGAT<br />
TGAAAAATAATTTTAACATTTTATATAATCAAATAAGACAGTACCCAGCTTATTATTTTAAAGTTGCTTCTAATGTTCCTACTTACTCTGA<br />
TATATGTCAATTTTTCTCAGTTATGTATCAAGGTTTTCAGATAGTTAATCATTCTGGTGATGTTTTTATTCATGCTTGTAGAGAAAATCCA<br />
CAATCTAAAGGTGATTTTGTTGGTGACAAGTTCCATATTTCTATAGCTAGAGAACAAGTTCCATTGGCATTTCAAATTTTATCTGGTTTGTT<br />
ATTTTCTGAGGACTCTCCTATTGACAAATGGAAGATAACTGATATGAATAGAGTTTCTCAACAGTCAAGAGTTGGTATTGGTGCTCAATTTAC<br />
TTTGTATGTTAAATCTGATCAAGAATGTTCTCAGTATTCAGCTTTATTGTTGCATAAAATTAGACAATTCATAATGTGCTTGGAATCTAATTT<br />
GTTAAGATCTAAGATTGCTCCAGGTGAATATCCTGCATCTGATGTTAGACCAGAGGATTGGAAGTACGTTTCATATAGAAATGAATTGAGATC<br />
TGACAGAGATGGTTCTGAAAGACAAGAGCAGATGTTAAGAGAAGAACCTTTTTATAGATTGATGATTGAG<br />
<br />
<br />
<br />
'''6-Adaptor bases, followed by “C” followed by another adaptor:'''<br />
GGTAGCGAT<br />
<br />
<br />
<br />
'''7-Zippers:'''<br />
For RR:<br />
GGTTCTGGTTCTGGTTCTGGTTCTCTGGAAATCCGTGCGGCGTTCCTGGAAAAAGAAAACACCGCGCTGCGTACCCGTGCGGCGGAACTGCG<br />
TAAACGTGTTGGTCGTTGCCGTAACATCGTTTCTAAATACGAAACCCGTTACGGTCCGCTG<br />
For EE:<br />
<br />
GGTTCCGGTAGCGGTTCCGGTTCTATCACTATTAGGGCTGCTTTTTTGGAAAAAGAAAATACTGCTTTGAGAACTGAAATTGCTGAATTGG<br />
AAAAAGAAGTTGGTAGATGTGAAAATATTGTTTCTAAATATGAAACTAGATATGGTCCATTG<br />
<br />
<br />
<br />
'''8-Final adaptors, followed by “D”, followed by BamHI, 3 STOPS in three different frames, then NotI, the Adh terminator and SacI:'''<br />
<br />
<br />
TAATGCGGGATCCTGAGTAAATAAGCGGCCGCGCGAATTTC<br />
TTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTAT<br />
ACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGT<br />
AACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT<br />
ATTGACCACACCGAGCTC<br />
<br />
<br />
<u>'''PROJECT 2</u><br />
<br />
<br />
(1) PspOMI//pSte2-Ste2//XhoI//GFP//XmaI//RR//STOP//BamHI//AdhTerminator<br />
<br />
GGGCCCATCCAATATCACCTGACCTTCATCTTTATGTTTTTCCACTGGTGTAATGTACTCTGTTGCATCATCAATAAGCAACTGGGACGCCAATAATTCATACCAACAATCTTTTCTTAATGAGTTGTTTACAAACCCCAGAGGCCCTAAACCTATGTACCTCAAAGCTCGGACATCTTTTTTCTCTAATAACTTTGATATAACTTTTCCTTTTTGTTCTTCTGCCTTTCCACTGTCACAACGAGAAAGTGATTCCAGATATGCGTTATAACCTCCCCTTGTCAACGCGTCTTTGTCGTGGCTACTCTGATTAGTATGAAATAATGACCTTAATGGCATCGTATTTTGTTAATTGGCTTGTTCTATGTCATTTGCTAAATTTTCTCTTTATTTCTTTACTTTGCCAATTCAAGATAATCTTCCCTTCCCAGAGAGAAAAAAGGAAAAATTTAGCTATGAAACCTCAATAAGCTTTTTAATACACCAAAGATTCAAGATAAGAGCATAGAACGAACTGTAGAATAGTCCGGATATGTTATCCAATGCCTGCCAAAATGCATTGTCACACGCTGTAGTGCTCGAATAGGTGTTGCAATCCGTCAATATACGTCTTGCTCTGTGGGTAAATGTCTCGTGCATTAAGACAGGCTAGTATAAACGAGAAGAAGTATCCTGCTTTGCAATGAAACAATAGTATCCGCTAAGAATTTAAGCAGGCCAACGTCCATACTGCTTAGGACCTGTGCCTGGCAAGTCGCAGATTGAAGTTTTTTCAACCATGTAAATTTCCTAATTGGGTAAGTACATGATGAAACACATATGAAGAAAAAAGCTTTCCTACATATTCAAGATTTTTTTCTGTGGGTGGAATACTATTTAAGGAGTGCTATTAGTATCTTATTTGACTTCAAAGCAATACGATACCTTTTCTTTTCACCTGCTCTGGCTATAATTATAATTGGTTACTTAAAAATGCACCGTTAAGAACCATATCCAAGAATCAAAAATGTCTGATGCGGCTCCTTCATTGAGCAATCTATTTTATGATCCAACGTATAATCCTGGTCAAAGCACCATTAACTACACTTCCATATATGGGAATGGATCTACCATCACTTTCGATGAGTTGCAAGGTTTAGTTAACAGTACTGTTACTCAGGCCATTATGTTTGGTGTCAGATGTGGTGCAGCTGCTTTGACTTTGATTGTCATGTGGATGACATCGAGAAGCAGAAAAACGCCGATTTTCATTATCAACCAAGTTTCATTGTTTTTAATCATTTTGCATTCTGCACTCTATTTTAAATATTTACTGTCTAATTACTCTTCAGTGACTTACGCTCTCACCGGATTTCCTCAGTTCATCAGTAGAGGTGACGTTCATGTTTATGGTGCTACAAATATAATTCAAGTCCTTCTTGTGGCTTCTATTGAGACTTCACTGGTGTTTCAGATAAAAGTTATTTTCACAGGCGACAACTTCAAAAGGATAGGTTTGATGCTGACGTCGATATCTTTCACTTTAGGGATTGCTACAGTTACCATGTATTTTGTAAGCGCTGTTAAAGGTATGATTGTGACTTATAATGATGTTAGTGCCACCCAAGATAAATACTTCAATGCATCCACAATTTTACTTGCATCCTCAATAAACTTTATGTCATTTGTCCTGGTAGTTAAATTGATTTTAGCTATTAGATCAAGAAGATTCCTTGGTCTCAAGCAGTTCGATAGTTTCCATATTTTACTCATAATGTCATGTCAATCTTTGTTGGTTCCATCGATAATATTCATCCTCGCATACAGTTTGGAACCAAACCAGGGAACAGATGTCTTGACTACTGTTGCAACATTACTTGCTGTATTGTCTTTACCATTATCATCAATGTGGGCCACGGCTGCTAATAATGCATCCAAAACAAACACAATTACTTCAGACTTTACAACATCCACAGATAGGTTTTATCCAGGCACGCTGTCTAGCTTTCAAACTGATAGTATCAACAACGATGCTAAAAGCAGTCTCAGAAGTAGATTATATGACCTATATCCTAGAAGGAAGGAAACAACATCGGATAAACATTCGGAAAGAACTTTTGTTTCTGAGACTGCAGATGATATAGAGAAAAATCAGTTTTATCAGTTGCCCACACCTACGAGTTCAAAAAATACTAGGATAGGACCGTTTGCTGATGCAAGTTACAAAGAGGGAGAAGTTGAACCCGTCGACATGTACACTCCCGATACGGCAGCTGATGAGGAAGCCAGAAAGTTCTGGACTGAAGATAATAATAATTTACTCGAGTCTAAAGGTGAAGAATTATTCACTGGTGTTGTCCCAATTTTGGTTGAATTAGATGGTGATGTTAATGGTCACAAATTTTCTGTCTCCGGTGAAGGTGAAGGTGATGCTACTTACGGTAAATTGACCTTAAAATTTATTTGTACTACTGGTAAATTGCCAGTTCCATGGCCAACCTTAGTCACTACTTTAACTTATGGTGTTCAATGTTTTTCTAGATACCCAGATCATATGAAACAACATGACTTTTTCAAGTCTGCCATGCCAGAAGGTTATGTTCAAGAAAGAACTATTTTTTTCAAAGATGACGGTAACTACAAGACCAGAGCTGAAGTCAAGTTTGAAGGTGATACCTTAGTTAATAGAATCGAATTAAAAGGTATTGATTTTAAAGAAGATGGTAACATTTTAGGTCACAAATTGGAATACAACTATAACTCTCACAATGTTTACATCATGGCTGACAAACAAAAGAATGGTATCAAAGTTAACTTCAAAATTAGACACAACATTGAAGATGGTTCTGTTCAATTAGCTGACCATTATCAACAAAATACTCCAATTGGTGATGGTCCAGTCTTGTTACCAGACAACCATTACTTATCCACTCAATCTaggTTATCCAAAGATCCAAACGAAAAGAGAGACCACATGGTCTTGTTAGAATTTGTTACTGCTGCTGGTATTACCCATGGTATGGATGAATTGTACAAACCCGGGggatctggatcgggatctGATCCAGATTTGGAAATTAGAGCTGCTTTTTTGAGACAAAGAAATACTGCTTTGAGAACTGAAGTTGCTGAATTGGAACAAGAAGTTCAAAGATTGGAAAATGAAGTTTCTCAATATGAAACTAGATATGGTCCATTGGGTGGTGGTAAATAAGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC<br />
<br />
<br />
(2) PspOMI//pGal1//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
TGTTATTGTGATAGTATCTCAGCGGGATTTGTGTAGATTCACCTACAGCCTTTAAACCAGCAAAGTGAATTACCGAATCAATTTTATATTCTTTGAAAACCTTTTCCAGACCTTTTCGGTCACACAAATCAACCTCATAGAAGGGAATGTGATGCTTGGTCAAGACCTCTAACCTGGCTACAGAATCATAAGTTGAATTCGACAGGTTATCAGCAACAACACAGTCATATCCATTCTCAATTAGCTCTACCACAGTGTGTGAACCAATGTATCCAGCACCACCTGTAACCAAAACAATTTTAGAAGTACTTTCACTTTGTAACTGAGCTGTCATTTATATTGAATTTTCAAAAATTCTTACTTTTTTTTTGGATGGACGCAAAGAAGTTTAATAATCATATTACATGGCATTACCACCATATACATATCCATATCTAATCTTACTTATATGTTGTGGAAATGTAAAGAGCCCCATTATCTTAGCCTAAAAAAACCTTCTCTTTGGAACTTTCAGTAATACGCTTAACTGCTCATTGCTATATTGAAGTACGGATTAGAAGCCGCCGAGCGGGCGACAGCCCTCCGACGGAAGACTCTCCTCCGTGCGTCCTCGTCTTCACCGGTCGCGTTCCTGAAACGCAGATGTGCCTCGCGCCGCACTGCTCCGAACAATAAAGATTCTACAATACTAGCTTTTATGGTTATGAAGAGGAAAAATTGGCAGTAACCTGGCCCCACAAACCTTCAAATTAACGAATCAAATTAACAACCATAGGATGATAATGCGATTAGTTTTTTAGCCTTATTTCTGGGGTAATTAATCAGCGAAGCGATGATTTTTGATCTATTAACAGATATATAAATGGAAAAGCTGCATAACCACTTTAACTAATACTTTCAACATTTTCAGTTTGTATTACTTCTTATTCAAATGTCATAAAAGTATCAACAAAAAATTGTTAATATACCTCTATACTTTAACGTCAAGGAGAAAAAACTATAATGCTCGAGGGAGCTgcttctgcatcaacttctaaatataattcacactccttggagaatgagtctattaagaggacgtctcgagatggagtcaatcgagatctcactgaggctgttcctcgacttccaggagaaacactaatcactgacaaagaagttatttacatatgtcctttcaatggccccattaagggaagagtttacatcacaaattatcgtctttatttaagaagtttggaaacggattcttctctaatacttgatgttcctctgggtgtgatctcgagaattgaaaaaatgggaggcgcgacaagtagaggagaaaattcctatggtctagatattacttgtaaagacatgagaaacctgaggttcgctttgaaacaggaaggccacagcagaagagatatgtttgagatcctcacgagatacgcgtttcccctggctcacagtctgccattatttgcatttttaaatgaagaaaagtttaacgtggatggatggacagtttacaatccagtggaagaatacaggaggcagggcttgcccaatcaccattggagaataacttttattaataagtgctatgagctctgtgacacttaccctgctcttttggtggttccgtatcgtgcctcagatgatgacctccggagagttgcaacttttaggtcccgaaatcgaattccagtgctgtcatggattcatccagaaaataagacggtcattgtgcgttgcagtcagcctcttgtcggtatgagtgggaaacgaaataaagatgatgagaaatatctcgatgttatcagggagactaataaacaaatttctaaactcaccatttatgatgcaagacccagcgtaaatgcagtggccaacaaggcaacaggaggaggatatgaaagtgatgatgcatatcataacgccgaacttttcttcttagacattcataatattcatgttatgcgggaatctttaaaaaaagtgaaggacattgtttatcctaatgtagaagaatctcattggttgtccagtttggagtctactcattggttagaacatatcaagctcgttttgacaggagccattcaagtagcagacaaagtttcttcagggaagagttcagtgcttgtgcattgcagtgacggatgggacaggactgctcagctgacatccttggccatgctgatgttggatagcttctataggagcattgaagggttcgaaatactggtacaaaaagaatggataagttttggacataaatttgcatctcgaataggtcatggtgataaaaaccacaccgatgctgaccgttctcctatttttctccagtttattgattgtgtgtggcaaatgtcaaaacagttccctacagcttttgaattcaatgaacaatttttgattataattttggatcatctgtatagttgccgatttggtactttcttattcaactgtgaatctgctcgagaaagacagaaggttacagaaaggactgtttctttatggtcactgataaacagtaataaagaaaaattcaaaaaccccttctatactaaagaaatcaatcgagttttatatccagttgccagtatgcgtcacttggaactctgggtgaattactacattagatggaaccccaggatcaagcaacaacagccgaatccagtggagcagcgttacatggagctcttagccttacgcgacgaatacataaagcggcttgaggaactgcagctcgccaactctgccaagctttctgatcccccaacttcaccttccagtccttcgcaaatgatgccccatgtgcaaactcacttcGGTAGTTCCCTAggttccggtagcggttccggttctatcactattagggctgcttttttggaaaaagaaaatactgctttgagaactgaaattgctgaattggaaaaagaagttggtagatgtgaaaatattgtttctaaatatgaaactagatatggtccattgTAATCGCGGATCCGCGAATTTCTTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTATACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGTAACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT ATTGACCACACC<br />
<br />
(3) pGal1//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but WMTM1 sequence<br />
<br />
(4) pSte5//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but Ste5 promoter sequence:<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCTCTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTTCCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAGAAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAGTAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCACATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTAACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGGCGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAAAGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAAAGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTAGCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAAATG<br />
<br />
(5) pSte5//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above.<br />
<br />
(6) pCyc//ATG//Xho//[A]//HMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above, but Cyc promoter sequence:<br />
<br />
ctcgagcagatccgccaggcgtgtatatatagcgtggatggccaggcaactttagtgctgacacatacaggcatatatatatgtgtgcgacgacacatgatcatatggcatgcatgtgctctgtatgtatataaaactcttgttttcttcttttctctaaatattctttccttatacattaggacctttgcagcataaattactatacttctatagacacacaaacacaaatacacacactaaattaataatg<br />
<br />
(7) pCyc//ATG//Xho//[A]//WMTM1//[B]//E34(I)//STOP//[D]-BamHI-AdhTerminator<br />
<br />
As above.<br />
<br />
(8) pCyc//ATG//XhoI//GFP//XmaI//(SerGlySerGlySer)ING2 PH domain//STOP//BamHI<br />
<br />
As above, but //(SerGlySerGlySer)ING2 PH domain sequence as follows:<br />
<br />
TCTGGATCAGGTTCAGCTTCTCCAGTTGAATTTGCTATTGATCCTAACGAACCTACATACTGCTTATGTAATCAAGTTTCTTATGGTGAAATGATTGGTTGCGATAACGAGCAGTGTCCAATAGAATGGTTTCATTTCTCTTGCGTTTCTTTAACATATAAACCTAAGGGAAAATGGTACTGTCCTAAATGTAGAGGTGATAATGAGAAAACTATGGATAAGTCTACTGAAAAGACAAAAAAAGATAGAAGATCAAGA</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/IGEM:UCSF/2007/Plasmids
IGEM:UCSF/2007/Plasmids
2007-10-26T19:57:22Z
<p>Sergio Peisajovich: </p>
<hr />
<div><u>'''PROJECT 1</u><br />
<br />
<u>'''Expected sequences</u><br />
<br />
''Below you have the sequences of the different parts in the right order, as well as the adaptor sequences between parts. You will need to arrange them accordingly to your particular constructs, so that you will have 1-2-3-4-5-6-7-8.''<br />
<br />
<br />
<br />
'''1-Promoter sequence (variable, depends on particular acceptor)'''<br />
<br />
<br />
pFig1<br />
<br />
TTGAATGATCAACCAAACGCCGATATAGTCACTATTGGTGATAAAATTAAAATGC<br />
AGCTATTGAGAACCCATCCTAACAACATTAAATTGTCTATTAATGGAATTGGTAA<br />
AGATGCCCCAACTTTCCAAGAATCTGCTTTGATTGCCGATAAGTTATTGAGTGTC<br />
ATGAAGGCCGGCACTTACCCAAAGATTTCCATTTTCTACAATGACCCAGTGTCTT<br />
CCCTATCTTTTGAACCATCTGAAAAACCGATCTTTAACGCCAAGACCATTGAACA<br />
ATCCCCATCATTCGGCAAATTTGAGATCGACACGGACGCAAACGTTCCAAGAGAT<br />
TTGTTTGAATATACTTTGGCTAACCAAATGTTGACAGCAATGGCTCAAGGTTATG<br />
CTGCTGAAATTTCCGCCAGAAGAAACGCTATGGATAACGCTTCCAAGAATGCCGG<br />
TGATATGATCAATCGTTACTCTATCTTGTACAACAGAACAAGACAAGCTGTCATT<br />
ACTAATGAACTGGTTGATATTATTACTGGTGCTTCCTCTTTGGGATGATAAAAAA<br />
ATCACCCTGCATTGCCTCTTTATTTGACGTTGTTTTTGTAGAACATGAAACGAAT<br />
TTTGACTTGATGAGACGAAGTATATATCCAAAGAATACCTTAAATAGAAAAGGAA<br />
AGATAATAAATACTAAACACTACTATATATTCAGGTAAAATACAAAAATTATAAC<br />
ATTTTTAAAACTTTTTTTTTTTGAAAGTCCTTCTCGCTTTAGGATTTTTTCCCAT<br />
TAAGATTATGATGGTTTCATGTATGTGTCAGTTAAAAAAAATATGGCTAAGTAGC<br />
AATGAAACGAACCAAGAAAATGAAAACAATATATAGTGCTGTTGAAATAACAAAG<br />
ACATTGGTATATATTTGTAAAATGTCTGTTAAATGTTTTTTATCTCAGGTTCTTG<br />
CTTGTCTTTGGTAGAAGAAATTATAGTAAACAAACAAACAAACAAACAAAAAAAA<br />
AAAAAAAAAA<br />
<br />
pAdh<br />
<br />
TTAAAACAAGAAGAGGGTTGACTACATCACGATGAGGGGGATCGAAGAAATGATG<br />
GTAAATGAAATAGGAAATCAAGGAGCATGAAGGCAAAAGACAAATATAAGGGTCG<br />
AACGAAAAATAAAGTGAAAAGTGTTGATATGATGTATTTGGCTTTGCGGCGCCGA<br />
AAAAACGAGTTTACGCAATTGCACAATCATGCTGACTCTGTGGCGGACCCGCGCT<br />
CTTGCCGGCCCGGCGATAACGCTGGGCGTGAGGCTGTGCCCGGCGGAGTTTTTTG<br />
CGCCTGCATTTTCCAAGGTTTACCCTGCGCTAAGGGGCGAGATTGGAGAAGCAAT<br />
AAGAATGCCGGTTGGGGTTGCGATGATGACGACCACGACAACTGGTGTCATTATT<br />
TAAGTTGCCGAAAGAACCTGAGTGCATTTGCAACATGAGTATACTAGAAGAATGA<br />
GCCAAGACTTGCGAGACGCGAGTTTGCCGGTGGTGCGAACAATAGAGCGACCATG<br />
ACCTTGAAGGTGAGACGCGCATAACCGCTAGAGTACTTTGAAGAGGAAACAGCAA<br />
TAGGGTTGCTACCAGTATAAATAGACAGGTACATACAACACTGGAAATGGTTGTC<br />
TGTTTGAGTACGCTTTCAATTCATTTGGGTGTGCACTTTATTATGTTACAATATG<br />
GAAGGGAACTTTACACTTCTCCTATGCACATATATTAATTAAAGTCCAATGCTAG<br />
TAGAGAAGGGGGGTAACACCCCTCCGCGCTCTTTTCCGATTTTTTTCTAAACCGT<br />
GGAATATTTCGGATATCCTTTTGTTGTTTCCGGGTGTACAATATGGACTTCCTCT<br />
TTTCTGGCAACCAAACCCATACATCGGGATTCCTATAATACCTTCGTTGGTCTCC<br />
CTAACATGTAGGTGGCGGAGGGGAGATATACAATAGAACAGATACCAGACAAGAC<br />
ATAATGGGCTAAACAAGACTACACCAATTACACTGCCTCATTGATGGTGGTACAT<br />
AACGAACTAATACTGTAGCCCTAGACTTGATAGCCATCATCATATCGAAGTTTCA<br />
CTACCCTTTTTCCATTTGCCATCTATTGAAGTAATAATAGGCGCATGCAACTTCT<br />
TTTCTTTTTTTTTCTTTTCTCTCTCCCCCGTTGTTGTCTCACCATATCCGCAATG<br />
ACAAAAAAATGATGGAAGACACTAAAGGAAAAAATTAACGACAAAGACAGCACCA<br />
ACAGATGTCGTTGTTCCAGAGCTGATGAGGGGTATCTCGAAGCACACGAAACTTT<br />
TTCCTTCCTTCATTCACGCACACTACTCTCTAATGAGCAACGGTATACGGCCTTC<br />
CTTCCAGTTACTTGAATTTGAAATAAAAAAAAGTTTGCTGTCTTGCTATCAAGTA<br />
TAAATAGACCTGCAATTATTAATCTTTTGTTTCCTCGTCATTGTTCTCGTTCCCT<br />
TTCTTCCTTGTTTCTTTTTCTGCACAATATTTCAAGCTATACCAAGCATACAATC<br />
AACTATCTCATATACA<br />
<br />
<br />
pCyc<br />
<br />
CAGATCCGCCAGGCGTGTATATATAGCGTGGATGGCCAGGCAACTTTAGTGCTGA<br />
CACATACAGGCATATATATATGTGTGCGACGACACATGATCATATGGCATGCATG<br />
TGCTCTGTATGTATATAAAACTCTTGTTTTCTTCTTTTCTCTAAATATTCTTTCC<br />
TTATACATTAGGACCTTTGCAGCATAAATTACTATACTTCTATAGACACACAAAC<br />
ACAAATACACACACTAAATTAATA<br />
<br />
<br />
pSte5<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCT<br />
CTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTT<br />
CCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAG<br />
AAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAG<br />
TAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCAC<br />
ATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTA<br />
ACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGG<br />
CGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAA<br />
AGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAA<br />
AGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTA<br />
GCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAA<br />
<br />
<br />
'''2-ATG-XhoI-“A” and 2 adaptor bases:'''<br />
ATGCTCGAGGGAGCT<br />
<br />
<br />
'''3-mCherry:'''<br />
<br />
GTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGGAGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACG<br />
AGTTCGAGATCGAGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCAAGGGTGGCCCCCTGCC<br />
CTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCAAGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAG<br />
CTGTCCTTCCCCGAGGGCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGGACTCCTCCCTGC<br />
AGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCCCCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTG<br />
GGAGGCCTCCTCCGAGCGGATGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACGGCGGCCAC<br />
TACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGCCCGGCGCCTACAACGTCAACATCAAGTTGGACATCA<br />
CCTCCCACAACGAGGACTACACCATCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGTACAAG <br />
<br />
<br />
<br />
'''4-Adaptor bases, followed by “B” and another adaptor base:'''<br />
GGTAGTTCCCTA <br />
<br />
<br />
<br />
'''5-Effectors'''<br />
<br />
For YopH:<br />
<br />
AACTTATCATTAAGCGATCTTCATCGTCAGGTATCTCGATTGGTGCAGCAAGAGAGCGGTGATTGTACCGGGAAATTAAGAGGTAACGT<br />
TGCTGCCAATAAAGAAACTACCTTTCAAGGTTTGACCATAGCCAGTGGAGCCAGAGAGTCAGAAAAAGTATTTGCTCAAACTGTACTAA<br />
GCCACGTAGCAAATGTTGTTCTAACTCAAGAAGATACCGCTAAGCTATTGCAAAGTACGGTAAAGCATAATTTGAATAATTATGACTTA<br />
AGAAGTGTCGGCAATGGTAATAGTGTACTTGTCAGTTTACGTAGTGACCAAATGACACTACAAGACGCCAAAGTGCTGTTGGAGGCCGC<br />
ATTGCGACAAGAGTCGGGAGCGAGGGGGCATGTATCATCTCATTCACATTCAGCCCTTCACGCACCGGGAACCCCGGTGCGTGAAGGAC<br />
TGCGTTCACATCTAGACCCCAGAACTCCACCGTTGCCACCGCGTGAACGACCACACACTTCTGGCCATCACGGGGCTGGCGAAGCCAGA<br />
GCCACCGCACCAAGCACTGTTTCTCCTTATGGCCCAGAAGCGCGCGCAGAACTCAGCAGCCGCCTCACCACATTGCGCAATACGCTGGC<br />
GCCAGCAACGAATGATCCGCGTTACTTACAAGCCTGCGGCGGTGAAAAGCTAAACCGATTTAGAGATATTCAATGCTGTCGGCAAACCG<br />
CAGTACGCGCCGATCTTAATGCCAATTACATCCAGGTCGGTAACACTCGTACCATAGCGTGCCAGTATCCGCTACAATCTCAACTTGAA<br />
AGCCATTTCCGTATGCTGGCAGAAAACCGAACGCCAGTGTTGGCTGTTTTAGCGTCCAGTTCTGAGATAGCCAATCAAAGATTCGGTAT<br />
GCCAGATTATTTCCGCCAGAGTGGTACCTATGGCAGTATCACTGTAGAGTCTAAAATGACTCAGCAAGTTGGTCTCGGTGACGGGATTA<br />
TGGCAGATATGTATACTTTAACGATTCGTGAAGCGGGTCAAAAAACAATCTCTGTTCCTGTGGTTCATGTTGGCAATTGGCCCGATCAG<br />
ACCGCAGTCAGCTCTGAAGTTACCAAGGCACTCGCTTCACTGGTAGATCAAACAGCAGAAACAAAACGCAATATGTATGAAAGCAAAGG<br />
AAGTTCAGCGGTAGGAGATGACTCCAAATTACGGCCGGTAATACATTGCCGTGCGGGTGTTGGCCGTACTGCGCAACTGATTGGCGCAA<br />
TGTGCATGAATGATAGTCGTAATAGTCAGTTAAGCGTAGAAGATATGGTCAGCCAAATGCGAGTACAAAGAAATGGTATTATGGTACAA<br />
AAAGATGAGCAACTTGATGTTCTGATTAAGTTGGCTGAAGGACAAGGGCGACCATTATTAAATAGC<br />
<br />
For YopJ:<br />
<br />
ATCGGACCAATATCACAAATAAATATCTCCGGTGGCTTATCAGAAAAAGAGACCAGTTCTTTAATCAGTAATGAAGAGCTTAAAAATAT<br />
CATAACACAGTTGGAAACTGATATATCGGATGGATCCTGGTTCCATAAAAATTATTCACGTATGGATGTAGAAGTCATGCCCGCATTGG<br />
TAATCCAGGCGAACAATAAATATCCGGAAATGAATCTTAATCTTGTTACATCTCCATTGGACCTTTCAATAGAAATAAAAAACGTCATA<br />
GAAAATGGAGTTAGATCTTCCCGCTTCATAATTAACATGGGGGAAGGTGGAATACATTTCAGTGTAATTGATTACAAACATATAAATGG<br />
GAAAACATCTCTGATATTGTTTGAACCAGCAAACTTTAACAGTATGGGGCCAGCGATGCTGGCAATAAGGACAAAAACGGCTATTGAAC<br />
GTTATCAATTACCTGATTGCCATTTCTCCATGGTGGAAATGGATATTCAGCGAAGCTCATCTGAATGTGGTATTTTTAGTTTGGCACTG<br />
GCAAAAAAACTTTACATCGAGAGAGATAGCCTGTTGAAAATACATGAAGATAATATAAAAGGTATATTAAGTGATGGTGAAAATCCTTT<br />
ACCCCACGATAAGTTGGACCCGTATCTCCCGGTAACTTTTTACAAACATACTCAAGGTAAAAAACGTCTTAATGAATATTTAAATACTA<br />
ACCCGCAGGGAGTTGGTACTGTTGTTAACAAAAAAAATGAAACCATCGTTAATAGATTTGATAACAATAAATCCATTGTAGATGGAAAG<br />
GATTATCAGTTTCGGTACATAAAAAGAGAATAGCTGAATATAAAACACTTCTCAAAGTA<br />
<br />
For OspF:<br />
<br />
CCAATTAAAAAGCCTTGTTTGAAATTGAATTTAGATTCTTTGAACGTTGTTAGATCAGAAATTCCACAAATGTTATCTGCTAATGAAAGAT<br />
TGAAAAATAATTTTAACATTTTATATAATCAAATAAGACAGTACCCAGCTTATTATTTTAAAGTTGCTTCTAATGTTCCTACTTACTCTGA<br />
TATATGTCAATTTTTCTCAGTTATGTATCAAGGTTTTCAGATAGTTAATCATTCTGGTGATGTTTTTATTCATGCTTGTAGAGAAAATCCA<br />
CAATCTAAAGGTGATTTTGTTGGTGACAAGTTCCATATTTCTATAGCTAGAGAACAAGTTCCATTGGCATTTCAAATTTTATCTGGTTTGTT<br />
ATTTTCTGAGGACTCTCCTATTGACAAATGGAAGATAACTGATATGAATAGAGTTTCTCAACAGTCAAGAGTTGGTATTGGTGCTCAATTTAC<br />
TTTGTATGTTAAATCTGATCAAGAATGTTCTCAGTATTCAGCTTTATTGTTGCATAAAATTAGACAATTCATAATGTGCTTGGAATCTAATTT<br />
GTTAAGATCTAAGATTGCTCCAGGTGAATATCCTGCATCTGATGTTAGACCAGAGGATTGGAAGTACGTTTCATATAGAAATGAATTGAGATC<br />
TGACAGAGATGGTTCTGAAAGACAAGAGCAGATGTTAAGAGAAGAACCTTTTTATAGATTGATGATTGAG<br />
<br />
<br />
<br />
'''6-Adaptor bases, followed by “C” followed by another adaptor:'''<br />
GGTAGCGAT<br />
<br />
<br />
<br />
'''7-Zippers:'''<br />
For RR:<br />
GGTTCTGGTTCTGGTTCTGGTTCTCTGGAAATCCGTGCGGCGTTCCTGGAAAAAGAAAACACCGCGCTGCGTACCCGTGCGGCGGAACTGCG<br />
TAAACGTGTTGGTCGTTGCCGTAACATCGTTTCTAAATACGAAACCCGTTACGGTCCGCTG<br />
For EE:<br />
<br />
GGTTCCGGTAGCGGTTCCGGTTCTATCACTATTAGGGCTGCTTTTTTGGAAAAAGAAAATACTGCTTTGAGAACTGAAATTGCTGAATTGG<br />
AAAAAGAAGTTGGTAGATGTGAAAATATTGTTTCTAAATATGAAACTAGATATGGTCCATTG<br />
<br />
<br />
<br />
'''8-Final adaptors, followed by “D”, followed by BamHI, 3 STOPS in three different frames, then NotI, the Adh terminator and SacI:'''<br />
<br />
<br />
TAATGCGGGATCCTGAGTAAATAAGCGGCCGCGCGAATTTC<br />
TTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTAT<br />
ACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGT<br />
AACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT<br />
ATTGACCACACCGAGCTC<br />
<br />
<br />
<u>'''PROJECT 2</u></div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/IGEM:UCSF/2007/Plasmids
IGEM:UCSF/2007/Plasmids
2007-10-26T19:56:15Z
<p>Sergio Peisajovich: </p>
<hr />
<div><u>'''PROJECT 1</u><br />
<br />
<u>'''Expected sequences</u><br />
<br />
''Below you have the sequences of the different parts in the right order, as well as the adaptor sequences between parts. You will need to arrange them accordingly to your particular constructs, so that you will have 1-2-3-4-5-6-7-8.''<br />
<br />
<br />
<br />
'''1-Promoter sequence (variable, depends on particular acceptor)'''<br />
<br />
<br />
pFig1<br />
<br />
TTGAATGATCAACCAAACGCCGATATAGTCACTATTGGTGATAAAATTAAAATGC<br />
AGCTATTGAGAACCCATCCTAACAACATTAAATTGTCTATTAATGGAATTGGTAA<br />
AGATGCCCCAACTTTCCAAGAATCTGCTTTGATTGCCGATAAGTTATTGAGTGTC<br />
ATGAAGGCCGGCACTTACCCAAAGATTTCCATTTTCTACAATGACCCAGTGTCTT<br />
CCCTATCTTTTGAACCATCTGAAAAACCGATCTTTAACGCCAAGACCATTGAACA<br />
ATCCCCATCATTCGGCAAATTTGAGATCGACACGGACGCAAACGTTCCAAGAGAT<br />
TTGTTTGAATATACTTTGGCTAACCAAATGTTGACAGCAATGGCTCAAGGTTATG<br />
CTGCTGAAATTTCCGCCAGAAGAAACGCTATGGATAACGCTTCCAAGAATGCCGG<br />
TGATATGATCAATCGTTACTCTATCTTGTACAACAGAACAAGACAAGCTGTCATT<br />
ACTAATGAACTGGTTGATATTATTACTGGTGCTTCCTCTTTGGGATGATAAAAAA<br />
ATCACCCTGCATTGCCTCTTTATTTGACGTTGTTTTTGTAGAACATGAAACGAAT<br />
TTTGACTTGATGAGACGAAGTATATATCCAAAGAATACCTTAAATAGAAAAGGAA<br />
AGATAATAAATACTAAACACTACTATATATTCAGGTAAAATACAAAAATTATAAC<br />
ATTTTTAAAACTTTTTTTTTTTGAAAGTCCTTCTCGCTTTAGGATTTTTTCCCAT<br />
TAAGATTATGATGGTTTCATGTATGTGTCAGTTAAAAAAAATATGGCTAAGTAGC<br />
AATGAAACGAACCAAGAAAATGAAAACAATATATAGTGCTGTTGAAATAACAAAG<br />
ACATTGGTATATATTTGTAAAATGTCTGTTAAATGTTTTTTATCTCAGGTTCTTG<br />
CTTGTCTTTGGTAGAAGAAATTATAGTAAACAAACAAACAAACAAACAAAAAAAA<br />
AAAAAAAAAA<br />
<br />
pAdh<br />
<br />
TTAAAACAAGAAGAGGGTTGACTACATCACGATGAGGGGGATCGAAGAAATGATG<br />
GTAAATGAAATAGGAAATCAAGGAGCATGAAGGCAAAAGACAAATATAAGGGTCG<br />
AACGAAAAATAAAGTGAAAAGTGTTGATATGATGTATTTGGCTTTGCGGCGCCGA<br />
AAAAACGAGTTTACGCAATTGCACAATCATGCTGACTCTGTGGCGGACCCGCGCT<br />
CTTGCCGGCCCGGCGATAACGCTGGGCGTGAGGCTGTGCCCGGCGGAGTTTTTTG<br />
CGCCTGCATTTTCCAAGGTTTACCCTGCGCTAAGGGGCGAGATTGGAGAAGCAAT<br />
AAGAATGCCGGTTGGGGTTGCGATGATGACGACCACGACAACTGGTGTCATTATT<br />
TAAGTTGCCGAAAGAACCTGAGTGCATTTGCAACATGAGTATACTAGAAGAATGA<br />
GCCAAGACTTGCGAGACGCGAGTTTGCCGGTGGTGCGAACAATAGAGCGACCATG<br />
ACCTTGAAGGTGAGACGCGCATAACCGCTAGAGTACTTTGAAGAGGAAACAGCAA<br />
TAGGGTTGCTACCAGTATAAATAGACAGGTACATACAACACTGGAAATGGTTGTC<br />
TGTTTGAGTACGCTTTCAATTCATTTGGGTGTGCACTTTATTATGTTACAATATG<br />
GAAGGGAACTTTACACTTCTCCTATGCACATATATTAATTAAAGTCCAATGCTAG<br />
TAGAGAAGGGGGGTAACACCCCTCCGCGCTCTTTTCCGATTTTTTTCTAAACCGT<br />
GGAATATTTCGGATATCCTTTTGTTGTTTCCGGGTGTACAATATGGACTTCCTCT<br />
TTTCTGGCAACCAAACCCATACATCGGGATTCCTATAATACCTTCGTTGGTCTCC<br />
CTAACATGTAGGTGGCGGAGGGGAGATATACAATAGAACAGATACCAGACAAGAC<br />
ATAATGGGCTAAACAAGACTACACCAATTACACTGCCTCATTGATGGTGGTACAT<br />
AACGAACTAATACTGTAGCCCTAGACTTGATAGCCATCATCATATCGAAGTTTCA<br />
CTACCCTTTTTCCATTTGCCATCTATTGAAGTAATAATAGGCGCATGCAACTTCT<br />
TTTCTTTTTTTTTCTTTTCTCTCTCCCCCGTTGTTGTCTCACCATATCCGCAATG<br />
ACAAAAAAATGATGGAAGACACTAAAGGAAAAAATTAACGACAAAGACAGCACCA<br />
ACAGATGTCGTTGTTCCAGAGCTGATGAGGGGTATCTCGAAGCACACGAAACTTT<br />
TTCCTTCCTTCATTCACGCACACTACTCTCTAATGAGCAACGGTATACGGCCTTC<br />
CTTCCAGTTACTTGAATTTGAAATAAAAAAAAGTTTGCTGTCTTGCTATCAAGTA<br />
TAAATAGACCTGCAATTATTAATCTTTTGTTTCCTCGTCATTGTTCTCGTTCCCT<br />
TTCTTCCTTGTTTCTTTTTCTGCACAATATTTCAAGCTATACCAAGCATACAATC<br />
AACTATCTCATATACA<br />
<br />
<br />
pCyc<br />
<br />
CAGATCCGCCAGGCGTGTATATATAGCGTGGATGGCCAGGCAACTTTAGTGCTGA<br />
CACATACAGGCATATATATATGTGTGCGACGACACATGATCATATGGCATGCATG<br />
TGCTCTGTATGTATATAAAACTCTTGTTTTCTTCTTTTCTCTAAATATTCTTTCC<br />
TTATACATTAGGACCTTTGCAGCATAAATTACTATACTTCTATAGACACACAAAC<br />
ACAAATACACACACTAAATTAATA<br />
<br />
<br />
pSte5<br />
<br />
CACGAAGTGACTGACAGAATACTGACATCAGCTGATTTCTCATAGAGCTGTTTCT<br />
CTGATAACACGTTGTTTGAACATCGACAAGATGAAAATCTAGAAGTATCAAGTTT<br />
CCTTTAAAGGGATATATAACAGATTCTAAAACTGACAGAAATATTTCGAGTGAAG<br />
AAGAAGCGTTAAATATTGGATCTTTCCGCAGTTCTACTCTGATACATTTTTGAAG<br />
TAGGAGAGTCATTTAGAAGGCGTATTGCTCAATAGTAGAAAGCAGGCCTGTGCAC<br />
ATGAATTAATTAAAAAATATAAAGGTAGTGATTAGACGACACATGTCCATAGGTA<br />
ACCTGTCATAATTTTGAACAATTTCCCTTCTTTTCTTTTTTTTTTTTGGGTGCGG<br />
CGATATGTAGCTTGTTAATTTACACATCATGTACTTTTCTGCATCAAAATATGAA<br />
AGGCGATAGTAGCTAAAGAAAATACCGAGAATTTCCTCGAAAAGTTGACGACAAA<br />
AGAAAGGCATAAAAAAGTAATTTGAAAATATTTTAAAACTGTTTTAACCCATCTA<br />
GCATCCGCGCTAAAAAAGGAAGATACAGGATACAGCGGAAACAACTTTTAA<br />
<br />
<br />
'''2-ATG-XhoI-“A” and 2 adaptor bases:'''<br />
ATGCTCGAGGGAGCT<br />
<br />
<br />
'''3-mCherry:'''<br />
<br />
GTGAGCAAGGGCGAGGAGGATAACATGGCCATCATCAAGGAGTTCATGCGCTTCAAGGTGCACATGGAGGGCTCCGTGAACGGCCACG<br />
AGTTCGAGATCGAGGGCGAGGGCGAGGGCCGCCCCTACGAGGGCACCCAGACCGCCAAGCTGAAGGTGACCAAGGGTGGCCCCCTGCC<br />
CTTCGCCTGGGACATCCTGTCCCCTCAGTTCATGTACGGCTCCAAGGCCTACGTGAAGCACCCCGCCGACATCCCCGACTACTTGAAG<br />
CTGTCCTTCCCCGAGGGCTTCAAGTGGGAGCGCGTGATGAACTTCGAGGACGGCGGCGTGGTGACCGTGACCCAGGACTCCTCCCTGC<br />
AGGACGGCGAGTTCATCTACAAGGTGAAGCTGCGCGGCACCAACTTCCCCTCCGACGGCCCCGTAATGCAGAAGAAGACCATGGGCTG<br />
GGAGGCCTCCTCCGAGCGGATGTACCCCGAGGACGGCGCCCTGAAGGGCGAGATCAAGCAGAGGCTGAAGCTGAAGGACGGCGGCCAC<br />
TACGACGCTGAGGTCAAGACCACCTACAAGGCCAAGAAGCCCGTGCAGCTGCCCGGCGCCTACAACGTCAACATCAAGTTGGACATCA<br />
CCTCCCACAACGAGGACTACACCATCGTGGAACAGTACGAACGCGCCGAGGGCCGCCACTCCACCGGCGGCATGGACGAGCTGTACAAG <br />
<br />
<br />
<br />
'''4-Adaptor bases, followed by “B” and another adaptor base:'''<br />
GGTAGTTCCCTA <br />
<br />
<br />
<br />
'''5-Effectors'''<br />
<br />
For YopH:<br />
<br />
AACTTATCATTAAGCGATCTTCATCGTCAGGTATCTCGATTGGTGCAGCAAGAGAGCGGTGATTGTACCGGGAAATTAAGAGGTAACGT<br />
TGCTGCCAATAAAGAAACTACCTTTCAAGGTTTGACCATAGCCAGTGGAGCCAGAGAGTCAGAAAAAGTATTTGCTCAAACTGTACTAA<br />
GCCACGTAGCAAATGTTGTTCTAACTCAAGAAGATACCGCTAAGCTATTGCAAAGTACGGTAAAGCATAATTTGAATAATTATGACTTA<br />
AGAAGTGTCGGCAATGGTAATAGTGTACTTGTCAGTTTACGTAGTGACCAAATGACACTACAAGACGCCAAAGTGCTGTTGGAGGCCGC<br />
ATTGCGACAAGAGTCGGGAGCGAGGGGGCATGTATCATCTCATTCACATTCAGCCCTTCACGCACCGGGAACCCCGGTGCGTGAAGGAC<br />
TGCGTTCACATCTAGACCCCAGAACTCCACCGTTGCCACCGCGTGAACGACCACACACTTCTGGCCATCACGGGGCTGGCGAAGCCAGA<br />
GCCACCGCACCAAGCACTGTTTCTCCTTATGGCCCAGAAGCGCGCGCAGAACTCAGCAGCCGCCTCACCACATTGCGCAATACGCTGGC<br />
GCCAGCAACGAATGATCCGCGTTACTTACAAGCCTGCGGCGGTGAAAAGCTAAACCGATTTAGAGATATTCAATGCTGTCGGCAAACCG<br />
CAGTACGCGCCGATCTTAATGCCAATTACATCCAGGTCGGTAACACTCGTACCATAGCGTGCCAGTATCCGCTACAATCTCAACTTGAA<br />
AGCCATTTCCGTATGCTGGCAGAAAACCGAACGCCAGTGTTGGCTGTTTTAGCGTCCAGTTCTGAGATAGCCAATCAAAGATTCGGTAT<br />
GCCAGATTATTTCCGCCAGAGTGGTACCTATGGCAGTATCACTGTAGAGTCTAAAATGACTCAGCAAGTTGGTCTCGGTGACGGGATTA<br />
TGGCAGATATGTATACTTTAACGATTCGTGAAGCGGGTCAAAAAACAATCTCTGTTCCTGTGGTTCATGTTGGCAATTGGCCCGATCAG<br />
ACCGCAGTCAGCTCTGAAGTTACCAAGGCACTCGCTTCACTGGTAGATCAAACAGCAGAAACAAAACGCAATATGTATGAAAGCAAAGG<br />
AAGTTCAGCGGTAGGAGATGACTCCAAATTACGGCCGGTAATACATTGCCGTGCGGGTGTTGGCCGTACTGCGCAACTGATTGGCGCAA<br />
TGTGCATGAATGATAGTCGTAATAGTCAGTTAAGCGTAGAAGATATGGTCAGCCAAATGCGAGTACAAAGAAATGGTATTATGGTACAA<br />
AAAGATGAGCAACTTGATGTTCTGATTAAGTTGGCTGAAGGACAAGGGCGACCATTATTAAATAGC<br />
<br />
For YopJ:<br />
<br />
ATCGGACCAATATCACAAATAAATATCTCCGGTGGCTTATCAGAAAAAGAGACCAGTTCTTTAATCAGTAATGAAGAGCTTAAAAATAT<br />
CATAACACAGTTGGAAACTGATATATCGGATGGATCCTGGTTCCATAAAAATTATTCACGTATGGATGTAGAAGTCATGCCCGCATTGG<br />
TAATCCAGGCGAACAATAAATATCCGGAAATGAATCTTAATCTTGTTACATCTCCATTGGACCTTTCAATAGAAATAAAAAACGTCATA<br />
GAAAATGGAGTTAGATCTTCCCGCTTCATAATTAACATGGGGGAAGGTGGAATACATTTCAGTGTAATTGATTACAAACATATAAATGG<br />
GAAAACATCTCTGATATTGTTTGAACCAGCAAACTTTAACAGTATGGGGCCAGCGATGCTGGCAATAAGGACAAAAACGGCTATTGAAC<br />
GTTATCAATTACCTGATTGCCATTTCTCCATGGTGGAAATGGATATTCAGCGAAGCTCATCTGAATGTGGTATTTTTAGTTTGGCACTG<br />
GCAAAAAAACTTTACATCGAGAGAGATAGCCTGTTGAAAATACATGAAGATAATATAAAAGGTATATTAAGTGATGGTGAAAATCCTTT<br />
ACCCCACGATAAGTTGGACCCGTATCTCCCGGTAACTTTTTACAAACATACTCAAGGTAAAAAACGTCTTAATGAATATTTAAATACTA<br />
ACCCGCAGGGAGTTGGTACTGTTGTTAACAAAAAAAATGAAACCATCGTTAATAGATTTGATAACAATAAATCCATTGTAGATGGAAAG<br />
GATTATCAGTTTCGGTACATAAAAAGAGAATAGCTGAATATAAAACACTTCTCAAAGTA<br />
<br />
For OspF:<br />
<br />
CCAATTAAAAAGCCTTGTTTGAAATTGAATTTAGATTCTTTGAACGTTGTTAGATCAGAAATTCCACAAATGTTATCTGCTAATGAAAGAT<br />
TGAAAAATAATTTTAACATTTTATATAATCAAATAAGACAGTACCCAGCTTATTATTTTAAAGTTGCTTCTAATGTTCCTACTTACTCTGA<br />
TATATGTCAATTTTTCTCAGTTATGTATCAAGGTTTTCAGATAGTTAATCATTCTGGTGATGTTTTTATTCATGCTTGTAGAGAAAATCCA<br />
CAATCTAAAGGTGATTTTGTTGGTGACAAGTTCCATATTTCTATAGCTAGAGAACAAGTTCCATTGGCATTTCAAATTTTATCTGGTTTGTT<br />
ATTTTCTGAGGACTCTCCTATTGACAAATGGAAGATAACTGATATGAATAGAGTTTCTCAACAGTCAAGAGTTGGTATTGGTGCTCAATTTAC<br />
TTTGTATGTTAAATCTGATCAAGAATGTTCTCAGTATTCAGCTTTATTGTTGCATAAAATTAGACAATTCATAATGTGCTTGGAATCTAATTT<br />
GTTAAGATCTAAGATTGCTCCAGGTGAATATCCTGCATCTGATGTTAGACCAGAGGATTGGAAGTACGTTTCATATAGAAATGAATTGAGATC<br />
TGACAGAGATGGTTCTGAAAGACAAGAGCAGATGTTAAGAGAAGAACCTTTTTATAGATTGATGATTGAG<br />
<br />
<br />
<br />
'''6-Adaptor bases, followed by “C” followed by another adaptor:'''<br />
GGTAGCGAT<br />
<br />
<br />
<br />
'''7-Zippers:'''<br />
For RR:<br />
GGTTCTGGTTCTGGTTCTGGTTCTCTGGAAATCCGTGCGGCGTTCCTGGAAAAAGAAAACACCGCGCTGCGTACCCGTGCGGCGGAACTGCG<br />
TAAACGTGTTGGTCGTTGCCGTAACATCGTTTCTAAATACGAAACCCGTTACGGTCCGCTG<br />
For EE:<br />
<br />
GGTTCCGGTAGCGGTTCCGGTTCTATCACTATTAGGGCTGCTTTTTTGGAAAAAGAAAATACTGCTTTGAGAACTGAAATTGCTGAATTGG<br />
AAAAAGAAGTTGGTAGATGTGAAAATATTGTTTCTAAATATGAAACTAGATATGGTCCATTG<br />
<br />
<br />
<br />
'''8-Final adaptors, followed by “D”, followed by BamHI, 3 STOPS in three different frames, then NotI, the Adh terminator and SacI:'''<br />
<br />
<br />
TAATGCGGGATCCTGAGTAAATAAGCGGCCGCGCGAATTTC<br />
TTATGATTTATGATTTTTATTATTAAATAAGTTATAAAAAAAAATAAGTGTAT<br />
ACAAATTTTAAAGTGACTCTTAGGTTTTAAAACGAAAATTCTTATTCTTGAGT<br />
AACTCTTTCCTGTGGTCAGGTTGCTTTCTCAGGTATAGCATGAGGTCGCTCTT<br />
ATTGACCACACCGAGCTC</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/Microscopy
UCSF/Microscopy
2007-10-26T19:54:57Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''4- Microscopy'''<br />
<br />
:Analysis of pathway-dependent Ste2 endocytosis by fluorescence microscopy was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then transferred to a Concavaline-A treated 96-well fluorescence plate. Individual wells were observed at 100x in a Nikon Eclipse T2000 microscope. Ste2 endocytosis was induced by addition of 1 μM α-factor (Zymo Research) to activate the pathway. Time-lapse experiments were done by taking pictures every 1.5 minutes, for a total of up to 2 hours.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/Flow_Cytometry2
UCSF/Flow Cytometry2
2007-10-26T19:54:43Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''3- Flow Cytometry'''<br />
<br />
:Analysis of pathway-dependent Ste2 induction by flow cytometry was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then treated with 1 μM α-factor (Zymo Research) to activate the pathway. At 60 min. aliquots of cultures were removed, treated with cycloheximide (5μg/mL), and dispensed into 96-well culture plates. Following incubation at room temperature for 1 hr in the dark to allow for GFP fluorophore maturation, plates containing treated cultures were analyzed with a BD LSR-II flow cytometer (BD Biosciences) using a high-throughput sampling module. 10,000 cells were counted for each reading, and GFP fluorescence was measured by exciting at 488 nm with a 100 mW Coherent Sapphire laser. Mean fluorescent values reported are the result of at least 3 independent samples.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF
UCSF
2007-10-26T19:54:21Z
<p>Sergio Peisajovich: /* Behind the Scenes */</p>
<hr />
<div><div id="mainpage"><br />
[[Image:logo6.png]]<br />
</div><br />
<br />
{| cell spacing=“1”<br />
|width="300px" bgcolor="gray" |<br />
<!-- PEOPLE HEADER with edit link--><br />
<h3><html><img src=http://openwetware.org/images/3/39/Icon_groups.png alt=People><br />
</html><font color=black>People<br />
<hr/><br />
<!-- END PEOPLE HEADER with edit link--><br />
===Students===<br />
*[[Michael Chen]] (UC Berkeley)<br />
*[[Eric Chou]] (Lincoln High)<br />
*[[Jimmy Huang]] (Lincoln High)<br />
*[[Lauren Jann]] (Lincoln High)<br />
*[[Eric Meltzer]] (Palo Alto High)<br />
*[[Alexander Ng]] (Lincoln High)<br />
*[[Robert Ovadia]] (Lincoln High)<br />
<br />
===Instructors===<br />
*[[Noah Helman]]<br />
*[[Andrew Horwitz]]<br />
*[[Sergio Peisajovich]]<br />
*[[David Pincus]]<br />
*[[Ryan Ritterson]]<br />
*[[Nili Sommovilla]]<br />
*[[Reid Williams]]<br />
*[[Angela Won]]<br />
<br />
===Faculty Advisors===<br />
*[http://www.ucsf.edu/limlab/ Wendell Lim]<br />
*[[Hana El-Samad]]<br />
*[http://kortemmelab.ucsf.edu/ Tanja Kortemme]<br />
*[http://www.voigtlab.ucsf.edu/ Chris Voigt] <br />
</font><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
|width="280px" bgcolor="gainsboro" |<br />
<!-- COMMUNITY HEADER with edit link--><br />
<html><h3><br />
<img src="http://openwetware.org/images/b/b9/Icon_board.png" alt=Resources><br />
</html>The Project</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
===About Us===<br />
*[[/Our Story/]]<br />
===A Guided Tour through the Project: JUDGES, THIS IS FOR YOU!===<br />
*[[/Project Description/]]<br />
*[[/Main Introduction/]]<br />
*'''Project One: Engineering Protein Complexes'''<br />
**[[/Intro/]]<br />
**[[/Results/]]<br />
**[[/Conclusions/]]<br />
*'''Project Two: The Synthetic Organelle'''<br />
**[[/Organelle Intro/| Intro]]<br />
**[[/Results_P2/| Results]]<br />
**[[/Conclusions_P2/| Conclusions]]<br />
*[[/Next Steps/]]<br />
<br />
===Behind the Scenes===<br />
*'''Methods and Materials'''<br />
**'''Project One'''<br />
***[[/Cloning Strategy1/| Cloning Strategy]]<br />
***[[/YeastStrains1/| Yeast Strains]]<br />
***[[/Flow Cytometry1/| Flow Cytometry]]<br />
**'''Project Two'''<br />
***[[/Cloning Strategy2/| Cloning Strategy]]<br />
***[[/YeastStrains2/|Yeast Strains]]<br />
***[[/Flow Cytometry2/|Flow Cytometry]]<br />
***[[/Microscopy/|Microscopy]]<br />
*[[IGEM:UCSF/2007/Plasmids | Plasmids & Sequences]]<br />
<br />
<br />
<br />
<br />
<br />
<br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
|width="300px" bgcolor="gray" | <br />
<!-- NEWS HEADER with edit link--><br />
<font color=black><h3><html><br />
<img src="http://openwetware.org/images/e/e2/Icon_info.png" alt="News" border=”0”><br />
</html>Information</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
<br />
===Planning & Development===<br />
<br />
*[[/Weekly Meetings/]]<br />
*[[/Brainstorming/]]<br />
*[[/Past Presentations/]]<br />
*[[/Extra Materials/]]<br />
*[[/Papers of Interest/]]<br />
<br />
===Media===<br />
*[[IGEM:UCSF/2007/Photos | Photos]]<br />
*[[/Videos/]]<br />
*[http://169.230.117.36/today/cache/feature/200708022.html in ucsf news!]<br />
<br />
===Science, Arts & Crafts===<br />
*[[Felt-mation]]<br />
*[[Lab Realia]]<br />
*[[Logo Galore...]]<br />
<br />
===iGEM links===<br />
Tools<br />
*[http://www.pubmed.org PubMed]<br />
*[http://www.yeastgenome.org Yeast DB]<br />
*[http://openwetware.org/wiki/Protocols OpenWetWare Protocols]<br />
For Visitors<br />
*[http://www.ucsf.edu UCSF Homepage]<br />
*[http://partsregistry.org Registry of Parts]<br />
*[https://2007.igem.org/wiki/index.cgi iGEM 2007]<br />
*Who's been checking us out?<br />
<html> <a href="http://www3.clustrmaps.com/counter/maps.php?url=http://openwetware.org/wiki/IGEM:UCSF/2007"><img src="http://www3.clustrmaps.com/stats/maps-no_clusters/openwetware.org-wiki-IGEM-UCSF-2007-thumb.jpg" /><br />
</a> </html></font><br />
<br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
__NOTOC__</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/Flow_Cytometry2
UCSF/Flow Cytometry2
2007-10-26T19:52:57Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''3- Flow Cytometry'''<br />
<br />
:Analysis of pathway-dependent Ste2 induction by flow cytometry was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then treated with 1 μM α-factor (Zymo Research) to activate the pathway. At 60 min. aliquots of cultures were removed, treated with cycloheximide (5μg/mL), and dispensed into 96-well culture plates. Following incubation at room temperature for 1 hr in the dark to allow for GFP fluorophore maturation, plates containing treated cultures were analyzed with a BD LSR-II flow cytometer (BD Biosciences) using a high-throughput sampling module. 10,000 cells were counted for each reading, and GFP fluorescence was measured by exciting at 488 nm with a 100 mW Coherent Sapphire laser. Mean fluorescent values reported are the result of at least 3 independent samples.<br />
<br />
<br />
'''Microscopy'''<br />
<br />
:Analysis of pathway-dependent Ste2 endocytosis by fluorescence microscopy was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then transferred to a Concavaline-A treated 96-well fluorescence plate. Individual wells were observed at 100x in a Nikon Eclipse T2000 microscope. Ste2 endocytosis was induced by addition of 1 μM α-factor (Zymo Research) to activate the pathway. Time-lapse experiments were done by taking pictures every 1.5 minutes, for a total of up to 2 hours.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/Flow_Cytometry2
UCSF/Flow Cytometry2
2007-10-26T19:52:40Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''3- Flow Cytometry'''<br />
<br />
:Analysis of pathway-dependent Ste2 induction by flow cytometry was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then treated with 1 μM α-factor (Zymo Research) to activate the pathway. At 60 min. aliquots of cultures were removed, treated with cycloheximide (5μg/mL), and dispensed into 96-well culture plates. Following incubation at room temperature for 1 hr in the dark to allow for GFP fluorophore maturation, plates containing treated cultures were analyzed with a BD LSR-II flow cytometer (BD Biosciences) using a high-throughput sampling module. 10,000 cells were counted for each reading, and GFP fluorescence was measured by exciting at 488 nm with a 100 mW Coherent Sapphire laser. Mean fluorescent values reported are the result of at least 3 independent samples.<br />
<br />
<br />
'''3- Flow Cytometry'''<br />
<br />
:Analysis of pathway-dependent Ste2 endocytosis by fluorescence microscopy was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then transferred to a Concavaline-A treated 96-well fluorescence plate. Individual wells were observed at 100x in a Nikon Eclipse T2000 microscope. Ste2 endocytosis was induced by addition of 1 μM α-factor (Zymo Research) to activate the pathway. Time-lapse experiments were done by taking pictures every 1.5 minutes, for a total of up to 2 hours.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF
UCSF
2007-10-24T23:02:27Z
<p>Sergio Peisajovich: /* A Guided Tour through the Project */</p>
<hr />
<div><div id="mainpage"><br />
[[Image:logo6.png]]<br />
</div><br />
<br />
{| cell spacing=“1”<br />
|width="300px" bgcolor="gainsboro" |<br />
<!-- PEOPLE HEADER with edit link--><br />
<h3><html><img src=http://openwetware.org/images/3/39/Icon_groups.png alt=People><br />
</html><font color=black>People<br />
<hr/><br />
<!-- END PEOPLE HEADER with edit link--><br />
===Students===<br />
*[[Michael Chen]] (UC Berkeley)<br />
*[[Eric Chou]] (Lincoln High)<br />
*[[Jimmy Huang]] (Lincoln High)<br />
*[[Lauren Jann]] (Lincoln High)<br />
*[[Eric Meltzer]] (Palo Alto High)<br />
*[[Alexander Ng]] (Lincoln High)<br />
*[[Robert Ovadia]] (Lincoln High)<br />
<br />
===Instructors===<br />
*[[Noah Helman]]<br />
*[[Andrew Horwitz]]<br />
*[[Sergio Peisajovich]]<br />
*[[David Pincus]]<br />
*[[Ryan Ritterson]]<br />
*[[Nili Sommovilla]]<br />
*[[Reid Williams]]<br />
*[[Angela Won]]<br />
<br />
===Faculty Advisors===<br />
*[http://www.ucsf.edu/limlab/ Wendell Lim]<br />
*[[Hana El-Samad]]<br />
*[http://kortemmelab.ucsf.edu/ Tanja Kortemme]<br />
*[http://www.voigtlab.ucsf.edu/ Chris Voigt] <br />
</font><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
|width="280px" bgcolor="gray" |<br />
<!-- COMMUNITY HEADER with edit link--><br />
<html><h3><br />
<img src="http://openwetware.org/images/b/b9/Icon_board.png" alt=Resources><br />
</html>The Project</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
===About Us===<br />
*[[/Our Story/]]<br />
===A Guided Tour through the Project: JUDGES, THIS IS FOR YOU!===<br />
*[[/Project Description/]]<br />
*[[/Main Introduction/]]<br />
*'''Project One: Engineering Protein Complexes'''<br />
**[[/Intro/]]<br />
**[[/Results/]]<br />
**[[/Conclusions/]]<br />
*'''Project Two: The Synthetic Organelle'''<br />
**[[/Organelle Intro/| Intro]]<br />
**[[/Results_P2/| Results]]<br />
**[[/Conclusions_P2/| Conclusions]]<br />
*[[/Next Steps/]]<br />
<br />
===Behind the Scenes===<br />
*'''Methods and Materials'''<br />
**'''Project One'''<br />
***[[/Cloning Strategy1/| Cloning Strategy]]<br />
***[[/YeastStrains1/| Yeast Strains]]<br />
***[[/Flow Cytometry1/| Flow Cytometry]]<br />
***[[/Modeling1/| Modeling]]<br />
**'''Project Two'''<br />
***[[/Cloning Strategy2/| Cloning Strategy]]<br />
***[[/YeastStrains2/|Yeast Strains]]<br />
***[[/Flow Cytometry2/|Flow Cytometry]]<br />
***[[/Modeling2/| Modeling]]<br />
*[[IGEM:UCSF/2007/Plasmids | Plasmids & Sequences]]<br />
<br />
<br />
<br />
<br />
<br />
<br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
|width="300px" bgcolor="lightgray" | <br />
<!-- NEWS HEADER with edit link--><br />
<font color=black><h3><html><br />
<img src="http://openwetware.org/images/e/e2/Icon_info.png" alt="News" border=”0”><br />
</html>Information</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
<br />
===Planning & Development===<br />
<br />
*[[/Weekly Meetings/]]<br />
*[[/Brainstorming/]]<br />
*[[/Past Presentations/]]<br />
*[[/Extra Materials/]]<br />
*[[/Papers of Interest/]]<br />
<br />
===Media===<br />
*[[IGEM:UCSF/2007/Photos | Photos]]<br />
*[[/Videos/]]<br />
*[http://169.230.117.36/today/cache/feature/200708022.html in ucsf news!]<br />
<br />
===Science, Arts & Crafts===<br />
*[[Felt-mation]]<br />
*[[Lab Realia]]<br />
*[[Logo Galore...]]<br />
<br />
===iGEM links===<br />
Tools<br />
*[http://www.pubmed.org PubMed]<br />
*[http://www.yeastgenome.org Yeast DB]<br />
*[http://openwetware.org/wiki/Protocols OpenWetWare Protocols]<br />
For Visitors<br />
*[http://www.ucsf.edu UCSF Homepage]<br />
*[http://partsregistry.org Registry of Parts]<br />
*[https://2007.igem.org/wiki/index.cgi iGEM 2007]<br />
*Who's been checking us out?<br />
<html> <a href="http://www3.clustrmaps.com/counter/maps.php?url=http://openwetware.org/wiki/IGEM:UCSF/2007"><img src="http://www3.clustrmaps.com/stats/maps-no_clusters/openwetware.org-wiki-IGEM-UCSF-2007-thumb.jpg" /><br />
</a> </html></font><br />
<br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
__NOTOC__</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF
UCSF
2007-10-24T23:01:29Z
<p>Sergio Peisajovich: /* Behind the Scenes: Our Experiments */</p>
<hr />
<div><div id="mainpage"><br />
[[Image:logo6.png]]<br />
</div><br />
<br />
{| cell spacing=“1”<br />
|width="300px" bgcolor="gainsboro" |<br />
<!-- PEOPLE HEADER with edit link--><br />
<h3><html><img src=http://openwetware.org/images/3/39/Icon_groups.png alt=People><br />
</html><font color=black>People<br />
<hr/><br />
<!-- END PEOPLE HEADER with edit link--><br />
===Students===<br />
*[[Michael Chen]] (UC Berkeley)<br />
*[[Eric Chou]] (Lincoln High)<br />
*[[Jimmy Huang]] (Lincoln High)<br />
*[[Lauren Jann]] (Lincoln High)<br />
*[[Eric Meltzer]] (Palo Alto High)<br />
*[[Alexander Ng]] (Lincoln High)<br />
*[[Robert Ovadia]] (Lincoln High)<br />
<br />
===Instructors===<br />
*[[Noah Helman]]<br />
*[[Andrew Horwitz]]<br />
*[[Sergio Peisajovich]]<br />
*[[David Pincus]]<br />
*[[Ryan Ritterson]]<br />
*[[Nili Sommovilla]]<br />
*[[Reid Williams]]<br />
*[[Angela Won]]<br />
<br />
===Faculty Advisors===<br />
*[http://www.ucsf.edu/limlab/ Wendell Lim]<br />
*[[Hana El-Samad]]<br />
*[http://kortemmelab.ucsf.edu/ Tanja Kortemme]<br />
*[http://www.voigtlab.ucsf.edu/ Chris Voigt] <br />
</font><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
|width="280px" bgcolor="gray" |<br />
<!-- COMMUNITY HEADER with edit link--><br />
<html><h3><br />
<img src="http://openwetware.org/images/b/b9/Icon_board.png" alt=Resources><br />
</html>The Project</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
===About Us===<br />
*[[/Our Story/]]<br />
===A Guided Tour through the Project===<br />
*[[/Project Description/]]<br />
*[[/Main Introduction/]]<br />
*'''Project One: Engineering Protein Complexes'''<br />
**[[/Intro/]]<br />
**[[/Results/]]<br />
**[[/Conclusions/]]<br />
*'''Project Two: The Synthetic Organelle'''<br />
**[[/Organelle Intro/| Intro]]<br />
**[[/Results_P2/| Results]]<br />
**[[/Conclusions_P2/| Conclusions]]<br />
*[[/Next Steps/]]<br />
===Behind the Scenes===<br />
*'''Methods and Materials'''<br />
**'''Project One'''<br />
***[[/Cloning Strategy1/| Cloning Strategy]]<br />
***[[/YeastStrains1/| Yeast Strains]]<br />
***[[/Flow Cytometry1/| Flow Cytometry]]<br />
***[[/Modeling1/| Modeling]]<br />
**'''Project Two'''<br />
***[[/Cloning Strategy2/| Cloning Strategy]]<br />
***[[/YeastStrains2/|Yeast Strains]]<br />
***[[/Flow Cytometry2/|Flow Cytometry]]<br />
***[[/Modeling2/| Modeling]]<br />
*[[IGEM:UCSF/2007/Plasmids | Plasmids & Sequences]]<br />
<br />
<br />
<br />
<br />
<br />
<br><br><br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
|width="300px" bgcolor="lightgray" | <br />
<!-- NEWS HEADER with edit link--><br />
<font color=black><h3><html><br />
<img src="http://openwetware.org/images/e/e2/Icon_info.png" alt="News" border=”0”><br />
</html>Information</h3><br />
<hr/><br />
<!-- END NEWS HEADER with edit link--><br />
<br />
===Planning & Development===<br />
<br />
*[[/Weekly Meetings/]]<br />
*[[/Brainstorming/]]<br />
*[[/Past Presentations/]]<br />
*[[/Extra Materials/]]<br />
*[[/Papers of Interest/]]<br />
<br />
===Media===<br />
*[[IGEM:UCSF/2007/Photos | Photos]]<br />
*[[/Videos/]]<br />
*[http://169.230.117.36/today/cache/feature/200708022.html in ucsf news!]<br />
<br />
===Science, Arts & Crafts===<br />
*[[Felt-mation]]<br />
*[[Lab Realia]]<br />
*[[Logo Galore...]]<br />
<br />
===iGEM links===<br />
Tools<br />
*[http://www.pubmed.org PubMed]<br />
*[http://www.yeastgenome.org Yeast DB]<br />
*[http://openwetware.org/wiki/Protocols OpenWetWare Protocols]<br />
For Visitors<br />
*[http://www.ucsf.edu UCSF Homepage]<br />
*[http://partsregistry.org Registry of Parts]<br />
*[https://2007.igem.org/wiki/index.cgi iGEM 2007]<br />
*Who's been checking us out?<br />
<html> <a href="http://www3.clustrmaps.com/counter/maps.php?url=http://openwetware.org/wiki/IGEM:UCSF/2007"><img src="http://www3.clustrmaps.com/stats/maps-no_clusters/openwetware.org-wiki-IGEM-UCSF-2007-thumb.jpg" /><br />
</a> </html></font><br />
<br><br><br><br><br><br><br><br><br />
<br />
<br />
<br />
__NOTOC__</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/Flow_Cytometry2
UCSF/Flow Cytometry2
2007-10-24T22:56:38Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''3- Flow Cytometry'''<br />
<br />
:Analysis of pathway-dependent Ste2 induction by flow cytometry was performed as follows: cultures were grown to early log phase (OD600=0.1-0.3) in complete synthetic dropout media and then treated with 1 μM α-factor (Zymo Research) to activate the pathway. At 60 min. aliquots of cultures were removed, treated with cycloheximide (5μg/mL), and dispensed into 96-well culture plates. Following incubation at room temperature for 1 hr in the dark to allow for GFP fluorophore maturation, plates containing treated cultures were analyzed with a BD LSR-II flow cytometer (BD Biosciences) using a high-throughput sampling module. 10,000 cells were counted for each reading, and GFP fluorescence was measured by exciting at 488 nm with a 100 mW Coherent Sapphire laser. Mean fluorescent values reported are the result of at least 3 independent samples.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/YeastStrains2
UCSF/YeastStrains2
2007-10-24T22:54:52Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''2- Yeast Strains'''<br />
<br />
:Saccharomyces cerevisiae strains: (1) SP163 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-RR, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2), (2) SP164 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-HMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2),(3) SP165 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-WMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2). <br />
<br />
:Subsequently, each pRS315-based construct was transformed (selection based on Leu2 marker) into SP163.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/YeastStrains2
UCSF/YeastStrains2
2007-10-24T22:54:36Z
<p>Sergio Peisajovich: </p>
<hr />
<div>'''2- Yeast Strains'''<br />
<br />
: 'Saccharomyces cerevisiae strains: (1) SP163 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-RR, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2), (2) SP164 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-HMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2),(3) SP165 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-WMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2). <br />
<br />
:Subsequently, each pRS315-based construct was transformed (selection based on Leu2 marker) into SP163.</div>
Sergio Peisajovich
http://2007.igem.org/wiki/index.php/UCSF/YeastStrains2
UCSF/YeastStrains2
2007-10-24T22:52:27Z
<p>Sergio Peisajovich: </p>
<hr />
<div>2- Yeast Strains<br />
<br />
'Saccharomyces cerevisiae strains: (1) SP163 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-RR, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2), (2) SP164 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-HMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2),(3) SP165 (W303 MATa, bar1::NatR, far1D, ste2D, ho:: pSte2-Ste2-GFP-WMTM1, mfa2::pCyc-mCherry-PH-ING2, his 3, trp1, leu2). <br />
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Subsequently, each pRS315-based construct was transformed (selection based on Leu2 marker) into SP163.</div>
Sergio Peisajovich