Difference between revisions of "Part:BBa K2762010"

Revision as of 08:28, 29 September 2018

PlacI-B0034-rbcX-B0034-rbcS-B0015

Usage

The rbcXS part encode the RbcS subunit of RubisCO and the chaperon RbcX. The RbcS affect the activity of RubisCO and RbcX contribute to the correct folding of the whole RubisCO enzyme.

Biology

The rbcX and rbcS gene are from cynobacteria Synechococcus elongatus PCC 7002. We designed the PlacI (BBa_R0010) promoter for the gene and cloned gene in DH5 alpha in order to test the function while consecutive expression.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 PlacI 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: BBa_K2762012.

Characterization

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 W3110. 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]

@@ Line 6: / Line 6: @@
 ===Biology===
-The <i>rbcX</i> and <i>rbcS</i> gene are from  cynobacteria Synechococcus elongatus PCC 7002. We designed the PlacI (BBa_R0010) promoter for the gene and cloned gene in DH5 alpha in order to test the function while consecutive expression.It has reported that the expression of RubisCO could not be detectable while only add single promoter of the <i>rbcL-rbcX-rbcS</i> gene fragment. Therefore we added another PlacI promoter on the upstream of <i>rbcX-rbcS</i> 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: BBa_K2762012.
+The <i>rbcX</i> and <i>rbcS</i> gene are from  cynobacteria <i>Synechococcus elongatus</i> PCC 7002. We designed the PlacI (BBa_R0010) promoter for the gene and cloned gene in DH5 alpha in order to test the function while consecutive expression.It has reported that the expression of RubisCO could not be detectable while only add single promoter of the <i>rbcL-rbcX-rbcS</i> gene fragment. Therefore we added another PlacI promoter on the upstream of <i>rbcX-rbcS</i> 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: BBa_K2762012.
 ===Characterization===
-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.
+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 W3110. We than did the SDS-PAGE to confirm the present of the RbcX and RbcS protein.