Difference between revisions of "Part:pSB1C3:Experience"

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<partinfo>pSB1C3 AddReview 5</partinfo>
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<I>Waterloo iGEM 2014</I>
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The pSB1C3 backbone has been made more versatile by turning it into a shuttle vector. By cloning a ''Staphylococcal'' selective marker (erythromycin resistance gene) and origin of replication, this part has been improved to be used in both ''E. coli'' and ''Staphylococcal'' organisms. The ''Staphylococcal'' parts have been incorporated into the backbone between the VR and the pMB1 replication origin in pSB1C3. The newly integrated parts have been sequence confirmed and a restriction digest of this part shows expected bands.
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[[File:Shuttle vector restriction digest.png]]
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The shuttle vector was successfully electroporated and maintained in ''Staphylococcus epidermidis'' and the newly transformed cell displayed erythromycin and chloramphenicol resistance.
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For more information, view the part page for this shuttle vector: <a = href="https://parts.igem.org/Part:BBa_K1323017">BBa_K1323017</a>
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Revision as of 12:13, 17 October 2014

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of pSB1C3

User Reviews

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Enter the review inofrmation here.

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Waterloo iGEM 2014

The pSB1C3 backbone has been made more versatile by turning it into a shuttle vector. By cloning a Staphylococcal selective marker (erythromycin resistance gene) and origin of replication, this part has been improved to be used in both E. coli and Staphylococcal organisms. The Staphylococcal parts have been incorporated into the backbone between the VR and the pMB1 replication origin in pSB1C3. The newly integrated parts have been sequence confirmed and a restriction digest of this part shows expected bands.

Shuttle vector restriction digest.png

The shuttle vector was successfully electroporated and maintained in Staphylococcus epidermidis and the newly transformed cell displayed erythromycin and chloramphenicol resistance.


For more information, view the part page for this shuttle vector: <a = href="https://parts.igem.org/Part:BBa_K1323017">BBa_K1323017</a>

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UNIPV-Pavia iGEM 2011

This cloning vector has been improved, cloning in it the strong pTet promoter, between E and X restriction sites; the mRFP coding sequence from BBa_J61002, has been placed between S and P restirction sites, in order to facilitate the cloning of coding sequences downstream pTet promoter in this high copy number plasmid.
For more details see the BBa_K516999 experience page.

The 2011 Nevada iGEM Team attempted to use pSB1C3 as a template to amplify the chloramphenicol resistance cassette for use in a new part. When they attempted to design primers for this purpose, we were unable to identify a start codon. We subsequently contacted Dr. Knight and Austin Che about the problem. They re-examined the sequence and found that the sequence in the registry was missing two nucleotides which resulted in an apparent frameshift. Upon fixing the sequence annotation, we were able to design our amplification primers for the chloramphenicol cassette.

iGEM JHU Wetware 2012 The pSB1C3 backbone was utilized to create a new plasmid backbone specifically engineered for use with the Golden Gate assembly method. Flanking regions were added within the prefix and suffix that facilitate assembly through the use of Bsa1 overhangs. This makes for a scarless ligation into the vector. The resulting sequence is a standard pSB1C3 with a part of choice, as the overhangs are removed during the Bsa1 cleavage. Three varieties of the backbone were made.

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