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Team:USTC/Project/protein/bkgrd
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Evelynzhang (Talk | contribs) (New page: A fusion protein is the product of joining two genes or two proteins /peptides together. This is achieved through the creation of a fusion gene which is done through the new standard enzym...) |
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In our experiments, the fusion proteins, which are used for the identification, localization and purification of BMC, consist of a RFP protein, a GFP protein, a GST protein or HIS protein. | In our experiments, the fusion proteins, which are used for the identification, localization and purification of BMC, consist of a RFP protein, a GFP protein, a GST protein or HIS protein. | ||
| - | First, the localization of targeted proteins to the shell protein assemblies was investigated by the fusion of RFP-PduA. We cloned the RFP gene to the | + | First, the localization of targeted proteins to the shell protein assemblies was investigated by the fusion of RFP-PduA. We cloned the RFP gene to the 5' end of pduA. When the fusion protein was produced, patches of red were observed throughout the cell. |
Second, we know that a N-terminal region of PduP is required for efficient packaging into the BMC. To test this, fusion proteins (P[1-14]-GFP, P[1-18]-GFP, P[1-64]-GFP) were used. Although P[1-14] and P[1-18] were also be taken into consideration, we created P[1-64]-GFP fusion protein in the end to study the transportation of BMC and examined subcellular localization of it by fluorescence and electron microscopy. | Second, we know that a N-terminal region of PduP is required for efficient packaging into the BMC. To test this, fusion proteins (P[1-14]-GFP, P[1-18]-GFP, P[1-64]-GFP) were used. Although P[1-14] and P[1-18] were also be taken into consideration, we created P[1-64]-GFP fusion protein in the end to study the transportation of BMC and examined subcellular localization of it by fluorescence and electron microscopy. | ||
| - | Third, a similar set of studies was done by fusing N-terminal sections of PduV to GFP. As it is reported, PduV localized to cup-like structures upon the outer surface of | + | Third, a similar set of studies was done by fusing N-terminal sections of PduV to GFP. As it is reported, PduV localized to cup-like structures upon the outer surface of BMCs. 98 amino acids from the N terminus of PduV were fused to GFP. Then we can observe the differential localization of PduV to the BMC. In addition, fusion proteins PduV-GST and PduV-HIS were expected to purify the BMC protein because of the affinity to special elution buffer. |
Revision as of 08:43, 26 October 2010
A fusion protein is the product of joining two genes or two proteins /peptides together. This is achieved through the creation of a fusion gene which is done through the new standard enzyme digestion of both the plasmids and the ligation of them. If it is two proteins that will be joined together, then a linker or spacer peptide will also be added due to our standard. This would usually make it more likely for the proteins to fold independently and behave as it should be.
In our experiments, the fusion proteins, which are used for the identification, localization and purification of BMC, consist of a RFP protein, a GFP protein, a GST protein or HIS protein.
First, the localization of targeted proteins to the shell protein assemblies was investigated by the fusion of RFP-PduA. We cloned the RFP gene to the 5' end of pduA. When the fusion protein was produced, patches of red were observed throughout the cell.
Second, we know that a N-terminal region of PduP is required for efficient packaging into the BMC. To test this, fusion proteins (P[1-14]-GFP, P[1-18]-GFP, P[1-64]-GFP) were used. Although P[1-14] and P[1-18] were also be taken into consideration, we created P[1-64]-GFP fusion protein in the end to study the transportation of BMC and examined subcellular localization of it by fluorescence and electron microscopy.
Third, a similar set of studies was done by fusing N-terminal sections of PduV to GFP. As it is reported, PduV localized to cup-like structures upon the outer surface of BMCs. 98 amino acids from the N terminus of PduV were fused to GFP. Then we can observe the differential localization of PduV to the BMC. In addition, fusion proteins PduV-GST and PduV-HIS were expected to purify the BMC protein because of the affinity to special elution buffer.
