Difference between revisions of "Part:BBa K1642009"

Latest revision as of 21:31, 18 September 2015

Expression of GFP under the control of Pdark

Figure1. Control mechanism of Pdark. (a) Under natural light, the green light component induces phosphorylation of CcaR, and the binding of it to Pdark prevents the binding of RNA polymerase, thus inhibiting expression of halorhodopsin. (b) Under darkness, RNA Polymerase binds to Pdark as it does in E.coli and initiate the transcription of halorhodopsin.

Pdark-GFP is a plasmid used to test whether the promoter Pdark works as we expect.

Up and Down are sequences uesd for homologous recombination.

Pdark(BBa_K1026009) is a “dark-sensing” promoter combines PcpcG2 and a constitutive promoter from E.coli. Pdark contains binding site of CcaR in PcpcG2, which overlaps with the binding site of RNA polymerase. Therefore when phosphorylated CcaR binds to it, the binding of RNA polymerase to Pdark will be blocked and transcription can’t be initiated.

RBS is the RNA binding site, which will hopefully increase the expression level of GFP.

PcpcB is a constitutive promoter in Synechocystis sp. strain PCC 6803, and KanaR is the gene sequence for kanamycin resistance, so that bacteria which are successfully transformed this plasmid can be screened on the plate.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 611
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 2995
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 481
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 1294

@@ Line 1: / Line 1: @@
 __NOTOC__
 <partinfo>BBa_K1642009 short</partinfo>
+[[File:SJTU Pdark.png|600px|thumb|left|Figure1. Control mechanism of Pdark. (a) Under natural light, the green light component induces phosphorylation of CcaR, and the binding of it to Pdark prevents the binding of RNA polymerase, thus inhibiting expression of halorhodopsin. (b) Under darkness, RNA Polymerase binds to Pdark as it does in E.coli and initiate the transcription of halorhodopsin. ]]
 Pdark-GFP is a plasmid used to test whether the promoter Pdark works as we expect.
@@ Line 6: / Line 8: @@
 Up and Down are sequences uesd for homologous recombination.
-Pdark(BBa_K1026009) is a “dark-sensing” promoter combines PcpcG2 and a constitutive promoter from ''E.coli''.Pdark contains binding site of CcaR in PcpcG2, which overlaps with the binding site of RNA polymerase. Therefore when phosphorylated CcaR binds to it, the binding of RNA polymerase to Pdark will be blocked and transcription can’t be initiated.
+Pdark(BBa_K1026009) is a “dark-sensing” promoter combines PcpcG2 and a constitutive promoter from ''E.coli''. Pdark contains binding site of CcaR in PcpcG2, which overlaps with the binding site of RNA polymerase. Therefore when phosphorylated CcaR binds to it, the binding of RNA polymerase to Pdark will be blocked and transcription can’t be initiated.
-[[File:SJTU Pdark.png|400px|thumb|left|Figure1. Control mechanism of Pdark. (a) Under natural light, the green light component? induces phosphorylation of CcaR, and the binding of it to Pdark prevent the binding of RNA polymerase, thus inhibiting expression of halorhodopsin. (b) Under darkness, RNA Polymerase binds to Pdark and initiate the transcription of halorhodopsin as usual. ]]
+RBS is the RNA binding site, which will hopefully increase the expression level of GFP.
+PcpcB is a constitutive promoter in ''Synechocystis sp''. strain PCC 6803, and KanaR is the gene sequence for kanamycin resistance, so that bacteria which are successfully transformed this plasmid can be screened on the plate.