Difference between revisions of "Part:BBa K300000"

Revision as of 17:54, 14 October 2010

BioBrick integrative base vector for E. coli

BBa_K300000 is an integrative base vector backbone which can be used to integrate the desired BioBrick system into the genome of E. coli. This base vector can specialized to target the desired integration site in the host genome.

How to propagate it before performing genome integration

The default version of this vector contains BBa_I52002, so it *must* be propagated in a ccdB-tolerant strain such as DB3.1 (BBa_V1005).

After the insertion of the desired BioBrick part in the cloning site, this vector does not contain a standard replication origin anymore, so it *must* be propagated in a pir+ or pir-116 strain such as BBa_K300984 or BBa_K300985 that can replicate the R6K conditional origin (BBa_J61001).

How to engineer it

How to perform genome integration

The integration into the E. coli chromosome can exploit the bacteriophage attP-mediated integration or the homologous recombination.

Detailed protocols about attP-mediated integration can be found here:

Anderson JC et al., 2010 (REFERENCE 1 [1])
Haldimann A and Wanner BL, 2001 (REFERENCE 8 [2])

Detailed protocols about homologous recombination can be found here:

Martinez-Morales F et al., 1999 (REFERENCE 11 [3])
Posfai G et al., 1997 (REFERENCE 12 [4])

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal XbaI site found at 169
Illegal XbaI site found at 1539
Illegal SpeI site found at 1727
12
INCOMPATIBLE WITH RFC[12]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 2150
Illegal NheI site found at 1558
Illegal NheI site found at 1974
Illegal SpeI site found at 2
Illegal SpeI site found at 1727
Illegal PstI site found at 16
Illegal NotI site found at 9
Illegal NotI site found at 2156
21
INCOMPATIBLE WITH RFC[21]
Plasmid lacks a prefix.
Plasmid lacks a suffix.
Illegal EcoRI site found at 2150
23
INCOMPATIBLE WITH RFC[23]
Illegal prefix found at 2150
Illegal suffix found at 2
Illegal XbaI site found at 169
Illegal XbaI site found at 1539
Illegal SpeI site found at 1727
25
INCOMPATIBLE WITH RFC[25]
Illegal prefix found at 2150
Plasmid lacks a suffix.
Illegal XbaI site found at 169
Illegal XbaI site found at 1539
Illegal XbaI site found at 2165
Illegal SpeI site found at 2
Illegal SpeI site found at 1727
Illegal PstI site found at 16
1000
INCOMPATIBLE WITH RFC[1000]
Plasmid lacks a prefix.
Plasmid lacks a suffix.

@@ Line 17: / Line 17: @@
 Detailed protocols about attP-mediated integration can be found here:
-*Anderson JC, Dueber JE, Leguia M, Wu GC, Goler JA, Arkin AP, Keasling JD (2010), BglBricks: A flexible standard for biological part assembly. Journal of Biological Engineering 2010 4:1.
+*Anderson JC et al., 2010 (REFERENCE 1 [https://parts.igem.org/Part:BBa_K300000:Design#References])
-*Haldimann A, Wanner BL (2001), Conditional-replication, integration, excision, and retrieval plasmid-host systems for gene structure-function studies of bacteria. Journal of Bacteriology, 183(21), p.6386-6393.
+*Haldimann A and Wanner BL, 2001 (REFERENCE 8 [https://parts.igem.org/Part:BBa_K300000:Design#References])
 Detailed protocols about homologous recombination can be found here:
-*Martinez-Morales F, Borges AC, Martinez A, Shanmaugam KT, Ingram LO (1999), Chromosomal Integration of Heterologous DNA in Escherichia coli with Precise Removal of Markers and Replicons Used during Construction. Journal of Bacteriology, Vol. 181, No. 22, November, p. 7143–7148.
+*Martinez-Morales F et al., 1999 (REFERENCE 11 [https://parts.igem.org/Part:BBa_K300000:Design#References])
-*Posfai G, Koob MD, Kirkpatrick HA, Blattner FR (1997), Versatile Insertion Plasmids for Targeted Genome Manipulations in Bacteria: Isolation, Deletion, and Rescue of the Pathogenicity Island LEE of the Escherichia coli O157:H7 Genome. Journal of Bacteriology, Vol. 179, No. 13July, p. 4426–4428.
+*Posfai G et al., 1997 (REFERENCE 12 [https://parts.igem.org/Part:BBa_K300000:Design#References])
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