Difference between revisions of "Part:BBa K1033207"

Revision as of 00:41, 4 October 2013 (view source) MikaelS (Talk | contribs) ← Older edit		Revision as of 00:42, 4 October 2013 (view source) MikaelS (Talk | contribs) (→‎Usage and Biology) Newer edit →
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	The backbone pSBLbE is a shuttle vector between E. coli, Lacotbacillus, and probably other lactic acid bacteria (LAB) like <i>Lactococcus lactis</i>. It has been used for subcloning in <i>E. coli</i> and to transform <i>Lactobacillus reuteri</i>.		The backbone pSBLbE is a shuttle vector between E. coli, Lacotbacillus, and probably other lactic acid bacteria (LAB) like <i>Lactococcus lactis</i>. It has been used for subcloning in <i>E. coli</i> and to transform <i>Lactobacillus reuteri</i>.
	===Usage and Biology===		===Usage and Biology===
−	Its replicon is known from litterature to replicate in a wide range of gram positive and gram negative species<sup>[[#Footnote 1|[1]]]</sup>, and the backbone is meant to be used for work in <i>E. coli</i> and different LAB, when it is very useful to do preliminary work in <i>E. coli</i>, and transfer finished constructs to LAB.	+	Its replicon is known from litterature to replicate in a wide range of gram positive and gram negative species<sup>[[#Footnote 1|[1]]]</sup>, and the backbone is meant to be used for work in <i>E. coli</i> and different LAB; when it is very useful to do preliminary work in <i>E. coli</i>, and transfer finished constructs to LAB.
	<i><b>Fig 1:</b></i>		<i><b>Fig 1:</b></i>
−
	=== Construction ===		=== Construction ===

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Revision as of 00:42, 4 October 2013

Shuttle vector pSBLbE for E. coli and Lactobacillus

The backbone pSBLbE is a shuttle vector between E. coli, Lacotbacillus, and probably other lactic acid bacteria (LAB) like Lactococcus lactis. It has been used for subcloning in E. coli and to transform Lactobacillus reuteri.

Usage and Biology

Its replicon is known from litterature to replicate in a wide range of gram positive and gram negative species^[1], and the backbone is meant to be used for work in E. coli and different LAB; when it is very useful to do preliminary work in E. coli, and transfer finished constructs to LAB.

Fig 1:

Construction

It was made by replacing the replicon of the BioBrick compatible plasmid pSB4C15 with a broad range replicon from the engineered plasmid pJP059. The replicon is also referred to as pSH71 and is related to pWV01 and the same family of rolling circle replicating plasmids.^[2]

We have also changed the resistance cassette to one conferring erythromycin resistance, taken from the reuteri plasmid [http://www.ncbi.nlm.nih.gov/nuccore/AY556392.1 pLUL631]. It is easy to change resistance again thanks to the extra cloning sites of pSB4C15.

Results

The vector has been verified to work in E. coli and to provide resistance against 250 µg/ml erythromycin on LB-agar plates (though our erythromycin is old, and may have lost some potency).

It has also been shown to grow in Lactobacillus reuteri 100-23 on MRS-plates containing 5 µg/ml erythromycin.

More coming soon, meanwhile, please see pSB4C15, which is very similar, for a bit more information.

References

^[1] I. Pérez-Arellano, M. Zúñiga, and G. Pérez-Martínez (2001), [http://www.sciencedirect.com/science/article/pii/S0147619X01915318 Construction of Compatible Wide-Host-Range Shuttle Vectors for Lactic Acid Bacteria and Escherichia coli], Plasmid 46 (2) 106-116. ^[2] I. Pérez-Arellano, M. Zúñiga, and G. Pérez-Martínez (2001), [http://www.sciencedirect.com/science/article/pii/S0147619X01915318 Construction of Compatible Wide-Host-Range Shuttle Vectors for Lactic Acid Bacteria and Escherichia coli], Plasmid 46 (2) 106-116.

Sequence and Features