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Present Challenges of
Experimenting With Plasmid Systems
Origin of replication (Ori) is a sequence where the plasmid replication begins. Plasmids with similar Ori sites use the same replication machinery.
Problem #1
Replication machinery does not discriminate between similar Ori sites, therefore smaller plasmids are replicated faster and rapidly outgrow larger plasmids, which leads to uneven plasmid ratios in a cell.
Problem #1
This is even more likely to occur when some plasmids are low copy number. When the cell divides, low copy plasmids tend to be lost in next generations.
Problem #1
This inability to stably co-maintain different plasmids over a number of generations in the same bacterial cell line is called plasmid incompatibility. One needs to use different types of Ori to avoid this trouble.
Problem #2
However, it often causes trouble due to limited Ori selection options or the lack of good combination of Ori and plasmid's copy number. Imagine you require a system consisting of two - low and high copy number plasmids. Try picking them from the list.
Problem #2
Currently no handy method of changing the copy number of particular plasmid exists. It must be either low, or medium, or high.
Problem #2
There is also no mechanism to adjust the copy number when so is required in any specific situation, for example: incoming cellular signals or special patterns.
Problem #2
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<img src="http://2017.igem.org/wiki/images/6/66/T--Vilnius-Lithuania--step-2-4-plasmid1.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/a/a6/T--Vilnius-Lithuania--step-2-4-plasmid2.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/3/39/T--Vilnius-Lithuania--step-2-4-plasmid3.png" class="plasmid" width="50" height="48"> <img src="http://2017.igem.org/wiki/images/6/66/T--Vilnius-Lithuania--step-2-4-plasmid1.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/a/a6/T--Vilnius-Lithuania--step-2-4-plasmid2.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/3/39/T--Vilnius-Lithuania--step-2-4-plasmid3.png" class="plasmid" width="50" height="48"> <img src="http://2017.igem.org/wiki/images/6/66/T--Vilnius-Lithuania--step-2-4-plasmid1.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/a/a6/T--Vilnius-Lithuania--step-2-4-plasmid2.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/3/39/T--Vilnius-Lithuania--step-2-4-plasmid3.png" class="plasmid" width="50" height="48"> <img src="http://2017.igem.org/wiki/images/6/66/T--Vilnius-Lithuania--step-2-4-plasmid1.png" class="plasmid" width="48" height="48"> <img src="http://2017.igem.org/wiki/images/4/42/T--Vilnius-Lithuania--step-2-2-dna.png" class="dna">
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