Difference between revisions of "Part:pSB1A10"
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<partinfo>pSB1A10 short</partinfo> | <partinfo>pSB1A10 short</partinfo> | ||
− | This plasmid can be used to do a rough characterization of the Input/Output curve for a PoPS-based device. Additionally, it can be used for screening of part or device libaries via FACS. More information can be [http://openwetware.org/index.php?title=Endy:Screening_plasmid_1.0 found here.] Note that this plasmid contains ccdb, so should be transformed into [[ | + | This plasmid can be used to do a rough characterization of the Input/Output curve for a PoPS-based device. Additionally, it can be used for screening of part or device libaries via FACS. More information can be [http://openwetware.org/index.php?title=Endy:Screening_plasmid_1.0 found here.] Note that this plasmid contains ccdb, so should be transformed into [[Part:BBa_V1005]], or another compatible strain for initial preparation. |
[[Image:ScreeningPlasmid1.0.PNG|600px|left|thumb|'''Schematic of Screening Plasmid 1.0:''' We are using the Pbad arabinose-inducible induction system [1] as a tunable input. GFP is a measure of input and RFP is a measure of output. A Biobricks cloning site enables easy insertion of any Biobricks part. RNase E sites create independence between the mRNA stability of the device being screened and the mRNA stability of the fluorescent proteins. In particular, we suspect mRFP1 contains internal RNaseE cut sites and have added a hairpin 5’ of the coding region to slow degradation by RNase E. [2] | [[Image:ScreeningPlasmid1.0.PNG|600px|left|thumb|'''Schematic of Screening Plasmid 1.0:''' We are using the Pbad arabinose-inducible induction system [1] as a tunable input. GFP is a measure of input and RFP is a measure of output. A Biobricks cloning site enables easy insertion of any Biobricks part. RNase E sites create independence between the mRNA stability of the device being screened and the mRNA stability of the fluorescent proteins. In particular, we suspect mRFP1 contains internal RNaseE cut sites and have added a hairpin 5’ of the coding region to slow degradation by RNase E. [2] |
Revision as of 15:20, 8 May 2007
Screening Plasmid v1.0
This plasmid can be used to do a rough characterization of the Input/Output curve for a PoPS-based device. Additionally, it can be used for screening of part or device libaries via FACS. More information can be [http://openwetware.org/index.php?title=Endy:Screening_plasmid_1.0 found here.] Note that this plasmid contains ccdb, so should be transformed into Part:BBa_V1005, or another compatible strain for initial preparation.
Note: The sequence below is incorrect because the backbone that the SP device was inserted into was psb1A2 rather than psb1A3, so there is a terminator (not annotated) in the sequence below at position 1044-1121 that isn't actually present in the DNA of psb1A10. Updating this will take some time because I need to change all the feature locations by hand - but the Vector NTI file is correct.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 5242
Illegal NheI site found at 4443
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 5248 - 21INCOMPATIBLE WITH RFC[21]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 5242
Illegal BglII site found at 89
Illegal BamHI site found at 4382
Illegal XhoI site found at 67 - 23INCOMPATIBLE WITH RFC[23]Illegal prefix found at 5242
Illegal suffix found at 2 - 25INCOMPATIBLE WITH RFC[25]Illegal prefix found at 5242
Plasmid lacks a suffix.
Illegal XbaI site found at 5257
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal AgeI site found at 674
Illegal AgeI site found at 786
Illegal AgeI site found at 4217 - 1000INCOMPATIBLE WITH RFC[1000]Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BsaI.rc site found at 2197
Illegal BsaI.rc site found at 5118
Illegal SapI site found at 4199
References
[1] Khlebnikov et al,Modulation of gene expression from the arabinose-inducible araBAD promoter. J Ind Microbiol Biotechnol. 2002 Jul;29(1):34-7.
[2] Effect of gene location, mRNA secondary structures, and RNase sites on expression of two genes in an engineered operon. Biotechnol Bioeng. 2002 Dec 30;80(7):762-76.