Difference between revisions of "Part:BBa K2762002"

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__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K2762002 short</partinfo>
 
<partinfo>BBa_K2762002 short</partinfo>
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===Backgriund===
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===Background===
 
The rbcS part encode the rbcS subunit of RubisCO. The rbcS affect the activity of RubisCO.
 
The rbcS part encode the rbcS subunit of RubisCO. The rbcS affect the activity of RubisCO.
  
 
===Mechanism===
 
===Mechanism===
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The rbcS genes are from cyanobacteria Synechococcus elongatus PCC7002. We designed the T7 promoter (BBa_I719005)for the gene and cloned gene in BL21  to ensure the high expression rate. It has reported that the expression of RubisCO could not be detectable while only add single promoter of the rbcL-rbcX-rbcS gene fragment. Therefore we added another T7 promoter on the upstream of rbcX-rbcS gene and named the part as rbcXS. We also did the coden optimization for the gene to ensure successful expression. The activation of RubisCo will be maximize after the binding of the RbcS subunit. To get more information, see our composite part: RubisCO.
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The rbcS genes are from cyanobacteria <i>Synechococcus elongatus</i> PCC7002. We designed the T7 promoter (BBa_I719005)for the gene and cloned gene in BL21  to ensure the high expression rate. It has reported that the expression of RubisCO could not be detectable while only add single promoter of the rbcL-rbcX-rbcS gene fragment. Therefore we added another T7 promoter on the upstream of rbcX-rbcS gene and named the part as rbcXS. We also did the coden optimization for the gene to ensure successful expression. The activation of RubisCo will be maximize after the binding of the RbcS subunit. To get more information, see our composite part: RubisCO.
  
 
===Expression in <i>E. coli</i>===
 
===Expression in <i>E. coli</i>===
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<partinfo>BBa_K2762002 parameters</partinfo>
 
<partinfo>BBa_K2762002 parameters</partinfo>
 
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===Source===
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Codon oprimized <i>Synechococcus elongatus</i> PCC7002.
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====References====
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[1] Janet Newman 1t and Steven Gutteridge2*. (1994,JUN.15). Structure of an effector-induced inactivated state of ribulose 1,5-bisphosphate carboxylase/oxygenase: the binary complex between enzyme and xylulose 1,5-bisphosphate.<i> Cell</i>
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[2] Inger Andersson <sup>a,</sup>*, Anders Backlund <sup>b</sup>. (2007, DEC.20). Structure and function of Rubisco. <i>Plant Physiology and Biochemistry</i>.
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[3] Fuyu Gong, Guoxia Liu, Xiaoyun Zhai,Jie Zhou, Zhen Cai and Yin Li1 .(2015,Jun 18). Quantitative analysis of an engineered CO<sub>2</sub>-fixing Escherichia coli reveals great potential of heterotrophic CO<sub>2</sub> fixation.<i> Biotechnology for Biofuels.</i>

Latest revision as of 14:08, 17 October 2018


Small subunit of the ribulose-bisphosphate carboxylase/oxygenase rbcS

Background

The rbcS part encode the rbcS subunit of RubisCO. The rbcS affect the activity of RubisCO.

Mechanism

The rbcS genes are from cyanobacteria Synechococcus elongatus PCC7002. We designed the T7 promoter (BBa_I719005)for the gene and cloned gene in BL21 to ensure the high expression rate. It has reported that the expression of RubisCO could not be detectable while only add single promoter of the rbcL-rbcX-rbcS gene fragment. Therefore we added another T7 promoter on the upstream of rbcX-rbcS gene and named the part as rbcXS. We also did the coden optimization for the gene to ensure successful expression. The activation of RubisCo will be maximize after the binding of the RbcS subunit. To get more information, see our composite part: RubisCO.

Expression in E. coli

We did two experiment to confirm the expression of this part. First we inserted the part on pSB1C3 and cloned the plasmid into DH5 alpha. We extracted the plasmid after the colony formed and did the enzyme digestion to confirm the insertion was successful. Second, we cloned the plasmid into BL21(DE3) and induced the promoter with IPTG. We than did the SDS PAGE to confirm the present of the RbcX and RbcS protein.


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
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Source

Codon oprimized Synechococcus elongatus PCC7002.

References

[1] Janet Newman 1t and Steven Gutteridge2*. (1994,JUN.15). Structure of an effector-induced inactivated state of ribulose 1,5-bisphosphate carboxylase/oxygenase: the binary complex between enzyme and xylulose 1,5-bisphosphate. Cell

[2] Inger Andersson a,*, Anders Backlund b. (2007, DEC.20). Structure and function of Rubisco. Plant Physiology and Biochemistry.

[3] Fuyu Gong, Guoxia Liu, Xiaoyun Zhai,Jie Zhou, Zhen Cai and Yin Li1 .(2015,Jun 18). Quantitative analysis of an engineered CO2-fixing Escherichia coli reveals great potential of heterotrophic CO2 fixation. Biotechnology for Biofuels.