Difference between revisions of "Part:BBa J63006"
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GPD promoter (''' [https://parts.igem.org/Part:BBa_K571001  BBa_K571001] ''') was used as a control.
 
GPD promoter (''' [https://parts.igem.org/Part:BBa_K571001  BBa_K571001] ''') was used as a control.
 
For the data, please refer to the GPD Promoter page (''' [https://parts.igem.org/Part:BBa_K571001  BBa_K571001] ''') and '''[http://2017.igem.org/Team:Kyoto/Results our Wiki Result]'''.
 
For the data, please refer to the GPD Promoter page (''' [https://parts.igem.org/Part:BBa_K571001  BBa_K571001] ''') and '''[http://2017.igem.org/Team:Kyoto/Results our Wiki Result]'''.
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===Contribution===
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<u>Addition of loop for hairpin-dsRNA to BBa_J63006</u>
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''' [http://2017.igem.org/Team:Kyoto  iGEM Kyoto 2017] ''' has modified this part so that it can also be used for dsRNA expression (see ''' [https://parts.igem.org/Part:BBa_K2403005  BBa_K2403005] ''').
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In order to kill pine wood nematodes by feeding RNAi, we used this part as a promoter to express hairpin-loop dsRNA in yeast. Then we confirmed the expression level of dsRNA in yeast by qRT-PCR and fed the yeast to pine wood nematodes.
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We attached a hairpin loop sequence behind this part so that you can easily make hairpin type dsRNA targeted to your favorite gene.
 +
You can create a plasmid transcribing dsRNA with a hairpin loop which can be expressed in budding yeast by cleaving with the restriction enzyme NotI, connecting the sense part between the promoter and loop parts, then cleaving with the restriction enzyme HindIII for ligating the antisense part.

Revision as of 06:57, 1 November 2017

yeast GAL1 promoter

GAL1 promoter from S. cerevisiae Contains Kozak sequence, part BBa_J63003.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 150
  • 1000
    COMPATIBLE WITH RFC[1000]


Promoter characterization

Team Czech Republic 2015 used this promoter very extensively. As a result, the characterization of this part was improved. See the Experience page for details.

Promoter Strength

JHU Igem 2011 is working on further characterization of this part through using GFP expression within the next month.


Promoter improvement

Team USTC 2016 modify this part to make it "ready to use". It is more convenient to be utilized now. See the Experience page for details.

Promoter dose =

Gal1 prevents growth in media containing galactose.

SCUT iGEM 2014 have designed a experiment to quantify Gal1 dose titer.


Figure 1.This year we have .



















Characterization by [http://2017.igem.org/Team:Kyoto iGEM Kyoto 2017]

We used this promoter in order to express long hairpin RNA in yeast. Long hairpin RNA targeting B. xylophilus AK1 mRNA or GFP mRNA was cloned downstream of the promoter and introduced into the 2-micron high copy number plasmid of budding yeast, and subsequently expressed in yeast. GPD promoter ( BBa_K571001 ) was used as a control. For the data, please refer to the GPD Promoter page ( BBa_K571001 ) and [http://2017.igem.org/Team:Kyoto/Results our Wiki Result].


Contribution

Addition of loop for hairpin-dsRNA to BBa_J63006 [http://2017.igem.org/Team:Kyoto iGEM Kyoto 2017] has modified this part so that it can also be used for dsRNA expression (see BBa_K2403005 ).

In order to kill pine wood nematodes by feeding RNAi, we used this part as a promoter to express hairpin-loop dsRNA in yeast. Then we confirmed the expression level of dsRNA in yeast by qRT-PCR and fed the yeast to pine wood nematodes.

We attached a hairpin loop sequence behind this part so that you can easily make hairpin type dsRNA targeted to your favorite gene. You can create a plasmid transcribing dsRNA with a hairpin loop which can be expressed in budding yeast by cleaving with the restriction enzyme NotI, connecting the sense part between the promoter and loop parts, then cleaving with the restriction enzyme HindIII for ligating the antisense part.