Difference between revisions of "Part:BBa K2665014"
 
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===Usage and Biology===
 
===Usage and Biology===
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SseNHX1 is the vacuolar Na(+)/H(+) antiporter. This antiporter enables to survive high osmotic pressure by sequestering excessive cytoplasmic Na(+) into vacuole.
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<p>
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The gene, SseNHX1 is created by DNA family shuffling;  Two homologous Na(+)/H(+) antiporters from halophytes Salicornia europaea (SeNHX1) and Suaeda salsa (SsNHX1) were shuffled.
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This is a Na+/H+ antiporter gene, which is a chimeric gene of two homologous Na+/H+ antiporters from halophytes Salicornia europaea (SeNHX1) and Suaeda salsa (SsNHX1). The Na+/H+ antiporter was obtained by DNA family shuffling.<br>
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By reference, SseNHX1 enables higher salt tolerance and sequester Na(+) than the two protein, SeNHX1 and SsNHX1.
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The study in 2015 has found that expression of this transporter in yeast mutant showed higher salt tolerance ability in yeast growth test than that of SsNHX1and SeNHX1. It also has founded that SseNHX1 protein in yeast cells accumulated more Na+ and slightly more K+ than the proteins of SsNHX1 and SeNHX1 did. [1]
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</p>
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==Characterization==
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[[File:T--Kyoto--SseNHX1.png|400px]] <br>
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[[File:T--Kyoto--cont.png|400px]]<br>
  
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Pictures shown above is colonies of<i> S. cerevisiae</i> &Delta;ENA1- strain on SD midium containing 400mM NaCl. In this spot assay, part BBa_K2665014 cloned to a yeast high-copy vector was used. <br>
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This result shows that SseNHX1 contributes to salt tolerance of yeasts.
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[[File:T--Kyoto--K_con.jpeg|400px]] <br>
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[[File:T--Kyoto--Na con.jpeg|400px]]<br>
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The graph shown above is K+ or Na+ concentration in cells of<i> S.cerevisiae</i> &Delta;ENA1&Delta;NHA1 strain. Each gene in the graph was cloned to yeast vectors, which were introduced to yeasts. The word “high” means high copy plasmid and the word “low” means low copy plasmids. These transformed yeasts are cultured in SD containing 400mM NaCl. Detailed data are on our wiki.'''[http://2018.igem.org/Team:Kyoto Kyoto2018]'''<br>
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These results show that SseNHX1 greatly contributes to accumulation of K+ and Na+ in a yeast cell.<br>
  
 
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<partinfo>BBa_K2665014 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2665014 SequenceAndFeatures</partinfo>
  
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===Reference===
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[1]G.Wu, G.Wang et al. (2015) A chimeric vacuolar Na+/H+antiporter gene evolved by DNA family shuffling confers increased salt tolerance in yeast, Journal of Biotechnology 203 1-8
  
 
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Latest revision as of 23:45, 17 October 2018


TDH3 promoter->SseNHX1->6xHis->CYC terminator

SseNHX1

Usage and Biology

This is a Na+/H+ antiporter gene, which is a chimeric gene of two homologous Na+/H+ antiporters from halophytes Salicornia europaea (SeNHX1) and Suaeda salsa (SsNHX1). The Na+/H+ antiporter was obtained by DNA family shuffling.
The study in 2015 has found that expression of this transporter in yeast mutant showed higher salt tolerance ability in yeast growth test than that of SsNHX1and SeNHX1. It also has founded that SseNHX1 protein in yeast cells accumulated more Na+ and slightly more K+ than the proteins of SsNHX1 and SeNHX1 did. [1]

Characterization

T--Kyoto--SseNHX1.png
T--Kyoto--cont.png

Pictures shown above is colonies of S. cerevisiae ΔENA1- strain on SD midium containing 400mM NaCl. In this spot assay, part BBa_K2665014 cloned to a yeast high-copy vector was used.
This result shows that SseNHX1 contributes to salt tolerance of yeasts.


T--Kyoto--K con.jpeg
T--Kyoto--Na con.jpeg
The graph shown above is K+ or Na+ concentration in cells of S.cerevisiae ΔENA1ΔNHA1 strain. Each gene in the graph was cloned to yeast vectors, which were introduced to yeasts. The word “high” means high copy plasmid and the word “low” means low copy plasmids. These transformed yeasts are cultured in SD containing 400mM NaCl. Detailed data are on our wiki.[http://2018.igem.org/Team:Kyoto Kyoto2018]
These results show that SseNHX1 greatly contributes to accumulation of K+ and Na+ in a yeast cell.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 2379
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Reference

[1]G.Wu, G.Wang et al. (2015) A chimeric vacuolar Na+/H+antiporter gene evolved by DNA family shuffling confers increased salt tolerance in yeast, Journal of Biotechnology 203 1-8