Difference between revisions of "Part:BBa J179000"

Revision as of 08:26, 20 December 2013

This part is an empty backbone vector for the usage in Bacillus subtilis. It integrates in the amyE locus and can be selected with Chloramphenicol (cat gene). It has a ampicillin resistance for cloning in E.coli. This backbone is a BioBricked version of the B. subtilis vector pDG1662 ([http://www.ncbi.nlm.nih.gov/pubmed/8973347 Guérout-Fleury et al.]).

This version is derived from BBa_K823023 and does not contain AgeI nor NgoMIV restriction sites, therefore is RFC25-compatible.

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

For handling of B. subtilis vectors, please see here.

The transformation into B. subtilis is explained here.

To check for integration of this vector into the amyE locus, the starch test can be used. The colonies obtained from the transformation into B.subtilis were streaked on an LB-Agar plate with 5 mg/L chloramphenicol and onto a starch plate (also W168 as a control) and incubated at 37°C over night. The starch plate is then covered with Lugol's iodine. Colonies without a bright surrounding are correct. We tested this vector with various inserts, for example the final construct of our Sporobeads with GFP. The starch test shows that the insertion in in the amyE locus and fluorescence microscopy reveals the functionality of the construct.

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 6084
Illegal SpeI site found at 2
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 6090
21
INCOMPATIBLE WITH RFC[21]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 6084
Illegal BamHI site found at 6066
Illegal XhoI site found at 3574
23
INCOMPATIBLE WITH RFC[23]
Illegal prefix found at 6084
Illegal suffix found at 2
25
INCOMPATIBLE WITH RFC[25]
Illegal prefix found at 6084
Plasmid lacks a suffix.
Illegal XbaI site found at 6099
Illegal SpeI site found at 2
Illegal PstI site found at 16
1000
INCOMPATIBLE WITH RFC[1000]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal BsaI.rc site found at 4501

@@ Line 1: / Line 1: @@
-This part is an empty backbone vector for the usage in ''Bacillus subtilis''. It integrates in the ''amyE'' locus and can be selected with Chloramphenicol (cat gene). It has a ampicillin resistance for cloning in ''E.coli''. This backbone is a ''BioBricked'' version of the ''B. subtilis'' vector pDG1662 ([http://www.ncbi.nlm.nih.gov/pubmed/8973347 Guérout-Fleury ''et al.''])
+This part is an empty backbone vector for the usage in ''Bacillus subtilis''. It integrates in the ''amyE'' locus and can be selected with Chloramphenicol (cat gene). It has a ampicillin resistance for cloning in ''E.coli''. This backbone is a ''BioBricked'' version of the ''B. subtilis'' vector pDG1662 ([http://www.ncbi.nlm.nih.gov/pubmed/8973347 Guérout-Fleury ''et al.'']).
-This version does not contain AgeI nor NgoMIV restriction sites, therefore is RFC25-compatible.
+This version is derived from [https://parts.igem.org/Part:BBa_K823023 BBa_K823023] and does not contain AgeI nor NgoMIV restriction sites, therefore is RFC25-compatible.
 <html> <a> <img src="https://static.igem.org/mediawiki/2012/e/e0/LMU-Munich-PSBBs1C.png" height=200"/></a></html>
@@ Line 19: / Line 19: @@
 We tested this vector with various inserts, for example the final construct of our Sporobeads with GFP. The starch test shows that the insertion in in the ''amy''E locus and fluorescence microscopy reveals the functionality of the construct.
-Also the iGEM-Team from [http://2012.igem.org/Team:Groningen/international_cooperation Groningen 2012] verified that this backbone works just fine.
 <br>
 <br>