Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Reshma Shetty
Date created: 2008-01-05 12:00:00
Date modified: 2015-08-31 04:07:42
ccdB and minimal pUC19 derived high copy origin
| Types | DnaRegion |
| Roles | plasmid Plasmid |
| Sequences | BBa_I52002_sequence (Version 1) |
Description
{||-
|width='35%'|
[[Image:CcdB_Plasmid.png]]
|width='65%'|
The CcdB protein, constitutively expressed by P1016, is lethal to most of the BioBrick cell strains, only [[Part:BBa_V1005|DB3.1]] is resistant.
|}
This part is designed to be located in the multiple cloning site of low copy BioBrick vectors. It is composed of two parts [[Part:BBa_P1016|BBa_P1016]] and [[Part:BBa_I50022|BBa_I50022]]. [[Part:BBa_P1016|BBa_P1016]] provides selection against non-insert containing clones (see below). [[Part:BBa_I50022|BBa_I50022]] is based on the high copy pUC19 origin. It enables the base vector to be propagated at high copy for ease of purification.
P1016 is used when putting BioBrick parts into BioBrick plasmids. The part to be inserted and the plasmid are cut with BioBrick enzymes and mixed. The mixture will include both the original uncut or religated plasmid and the desired structure. However, because of CcdB, all of the cells containing the original plasmid die and the surviving colonies are the desired result. I50020 is useful because it is a high copy origin located within the BioBricks cloning site. Any plasmids containing this part will be propagated (and thus purified) at high copy. Once the insert of interest is cloned into the plasmid, the I50022 part is eliminated and the plasmid returns to the copy number of the primary origin in the plasmid (usually low copy).
A new set of low copy BioBrick [[Help:Plasmids|plasmids]] are available with the I52001 insert. See I52001 Physical DNA for the current list of plasmids that are available in this form.
Notes
This part is a composite part of BBa_P1016 and BBa_I50022. It is designed to be placed in the multiple cloning site of BioBricks vectors. Vectors with this part in the multiple cloning site will be selected against during BioBricks assembly because BBa_P1016 is lethal to most cell strains. Also, BBa_I50022 enables the vector to be propagated at high copy regardless of the other origins on the vector.Source
ccdB and pUC19| Access | Instance | Definition |
| public public | BBa_P1016 BBa_I50022 | ccdB pUC19 ori |