Difference between revisions of "Part:BBa K823021"

Revision as of 16:59, 23 September 2012

pSB_Bs1C-lacZ (lacZ reporter vector for B. subtilis)

This part is a reporter vector for Bacillus subtilis. It integrates at the amyE locus. It has a ampicillin resistance for cloning in E.coli and chloramphenicol resistance for B.subtilis. The multiple cloning site is followed by a lacZ gene for reporter assays. This backbone is a BioBricked version of the B. subtilis vector pAC6.

This BioBrick is part of the LMU 2012 igem project [http://2012.igem.org/Team:LMU-Munich/Bacillus_BioBricks BacillusBioBrickBox]

Reference: [http://www.ncbi.nlm.nih.gov/pubmed/11902727 Stülke et al.]

Fig. 2: β-galactosidase assay and growth curve of strains carrying the promoters P_liaG (black) and P_veg (grey) fused to lacZ. β-galactosidase activity (Miller Units)and the growth curve values are the average of two independant clones with their standard deviation that were measured during the same experiment. Experiment shows representative data which was obtained in the same way from three independent experiments.

The two constitutive promoters P_liaG and P_veg were evaluated with the reporter vector pSBBs1C-lacZ which contains the lacZ reporter gene (Fig.2). Promoter activity leads to the expression of the β-galactosidase which directly correlates to the promoter activity. The β-galactosidase assay of the constitutive Bacillus promoters P_veg and P_liaG was repeated three times. Data show one representative result. In the beginning of the growth curve both promoters show only low activity. But then it increases to a maximum before it decreases to the begininng level after about seven hours (Data not shown). Summing up the course of activity of both promoters P_veg and P_liaG is very similar based on the growth curve. The highest β-galactosidase activity and therefore the highest activity of the promoter P_veg with a maximum of 65 Miller units can be found during the transition from the logarithmic to the stationary phase. This is about five times higher than the acitivity of the promoter P_liaG with a maximum activity of about 12 Miller Units.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 9771
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 9777
21
INCOMPATIBLE WITH RFC[21]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 9771
Illegal BglII site found at 8313
Illegal BamHI site found at 22
23
INCOMPATIBLE WITH RFC[23]
Illegal prefix found at 9771
Illegal suffix found at 2
25
INCOMPATIBLE WITH RFC[25]
Illegal prefix found at 9771
Plasmid lacks a suffix.
Illegal XbaI site found at 9786
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NgoMIV site found at 4562
Illegal NgoMIV site found at 6888
Illegal NgoMIV site found at 8688
1000
INCOMPATIBLE WITH RFC[1000]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BsaI site found at 4738
Illegal SapI.rc site found at 5820

@@ Line 4: / Line 4: @@
 This part is a reporter vector for Bacillus subtilis. It integrates at the amyE locus. It has a ampicillin resistance for cloning in E.coli and chloramphenicol resistance for B.subtilis. The multiple cloning site is followed by a lacZ gene for reporter assays. This backbone is a ''BioBricked'' version of the ''B. subtilis'' vector pAC6.
-<html> <a> <img src="https://static.igem.org/mediawiki/2012/0/04/LMU-Munich-PSBBs1C-lacZ.png" height=200"/></a></html>
+[[Image:LMU-Munich-PSBBs1C-lacZ.png|600px]]
 This BioBrick is part of the LMU 2012 igem project [http://2012.igem.org/Team:LMU-Munich/Bacillus_BioBricks '''''Bacillus''B'''io'''B'''rick'''B'''ox]
@@ Line 10: / Line 10: @@
 Reference: [http://www.ncbi.nlm.nih.gov/pubmed/11902727 Stülke ''et al''.]
+[[File:Englisch_Auswertung_PliaG_Pveg.png‎|thumb|right|400px| <p align="justify"> '''Fig. 2: β-galactosidase assay and growth curve of strains carrying the promoters P<sub>''liaG''</sub> (black) and P<sub>''veg''</sub> (grey) fused to ''lacZ'''''. β-galactosidase activity (Miller Units)and the growth curve values are the average of two independant clones with their standard deviation that were measured during the same experiment. Experiment shows representative data which was obtained in the same way from three independent experiments.</p>]]
+<p align="justify"> The two constitutive promoters P<sub>''liaG''</sub> and P<sub>''veg''</sub> were evaluated with the reporter vector pSBBs1C-lacZ which contains the lacZ reporter gene ('''Fig.2'''). Promoter activity leads to the expression of the β-galactosidase which directly correlates to the promoter activity. The β-galactosidase assay of the constitutive ''Bacillus'' promoters P<sub>''veg''</sub> and P<sub>''liaG''</sub> was repeated three times. Data show one representative result. In the beginning of the growth curve both promoters show only low activity. But then it increases to a maximum before it decreases to the begininng level after about seven hours (Data not shown). Summing up the course of activity of both promoters P<sub>''veg''</sub> and P<sub>''liaG''</sub> is very similar based on the growth curve. The highest β-galactosidase activity and therefore the highest activity of the promoter P<sub>''veg''</sub> with a maximum of 65 Miller units can be found during the transition from the logarithmic to the stationary phase. This is about five times higher than the acitivity of the promoter P<sub>''liaG''</sub> with a maximum activity of about 12 Miller Units.
+</p>
 <!-- Add more about the biology of this part here