Part:BBa_K5425002
This part encodes an antisense mRNA that is complementary to approximately 400 bp of the primary transcript of dnaA (a replication initiator protein) in Escherichia coli. The expression of this part downregulates the primary dnaA transcripts, ultimately lowering dnaA protein levels. This reduction is expected to lead to cell cycle arrest in E. coli.
Short description
This part encodes an antisense mRNA that is complementary to approximately 400 bp of the primary transcript of dnaA (a replication initiator protein) in Escherichia coli. The expression of this part downregulates the primary dnaA transcripts, ultimately lowering dnaA protein levels. This reduction is expected to lead to cell cycle arrest in E. coli.
General introduction to usage and biology
400 bp Antisense Characterization
The 400 bp antisense mRNA was the main antisense which was used for the quorum sensing circuit to induce cell cycle arrest. We hereby present the drylab characterization of the 400 bp antisense mRNA.
Fig: Secondary Structure and MFE for the 400 bp antisense mRNA
Fig: Bound structure of the 400 bp antisense mRNA with the sense mRNA of DnaA protein
Fig: ΔG of interaction vs length for antisense mRNAs of different lengths (from IntaRNA data)
Sense-Antisense Interaction Energy for the 400 bp antisense = –611 kcal/mol
Fig: Length based Half lives of the antisense mRNAs (data from DegScore)
Half life of the 400 bp antisense mRNA = 0.5776 hours.
We incorporated the Interaction Energy and Half Life for the 400 bp antisense mRNA obtained into our QS math model and determined the Bacterial population vs time :-
Fig: Bacterial Concentration vs time for the 400 bp antisense mRNA
Fig: Heat Map of the suitability of each of the antisense mRNAs based upon their lengths for achieving our desired target with the QS Circuit, as predicted by our software AntiRNA
The 400 bp antisense mRNA is in the dark blue region of the heat map, clearly denoting that the 400 bp antisense is well suitable in our QS circuit for achieving our desired target of controlling the bacterial population at the required threshold level quite below the carrying capacity for increasing the longevity of the bacterial population in the medium, much longer that what a general bacterial population in a medium usually lasts, by ensuring a slower rate of consumption of substrate as well as slower accumulation of secondary metabolites.
Sequence and Features
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal suffix found in sequence at 417
Illegal EcoRI site found at 1 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 1
Illegal SpeI site found at 418
Illegal PstI site found at 432
Illegal NotI site found at 7
Illegal NotI site found at 425 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 1
- 23INCOMPATIBLE WITH RFC[23]Illegal suffix found in sequence at 418
Illegal EcoRI site found at 1 - 25INCOMPATIBLE WITH RFC[25]Illegal prefix found in sequence at 1
Illegal SpeI site found at 418
Illegal PstI site found at 432 - 1000COMPATIBLE WITH RFC[1000]
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