15 Cloning Vectors
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• Alternative sequences which might be used for selection, see below.
• Optional (phage) promotors flanking the insertion site of the insert and
allowing in vitro transcription of cloned genes.
To keep plasmids stable (i.e., to prevent lass of part or all of the insert) and
to assure efficient transformation into bacterial cells, an insert length of
5000 bp should not be exceeded.
• pBR322 (Bolivar 1977, Sutcliffe 1978)
A double-stranded DNA plasmid cloning vehicle of about 4.4 kb in
length. pBR322 carries an origin of replication and resistance genes
for ampicillin and tetracycline. Several unique cloning sites within the
resistance genes allow for simple differentiation between plasmids bearing insertsandwild-type plasmids.
• pUC (Vieira 1982)
A double-stranded DNA plasmid cloning vehicle of about 2.7 kb in
length. pUC plasmids share in common with pBR322 the ampicillin resistance gene and the origin of replication. In addition they share in common with the M13 derived vectors the lacZ a-peptide gene of E. coli and
its multiple cloning region (MCR) (see below). This permits color-selection between plasmids bearing inserts and wild-type plasmids. As with
M13, pUC plasmids are available in pairs with reversed order of restriction sites in the MCR.
origin of
replication
T7 -promotor
ampicillinresistance-gene
plasmid
Bluescript
colour-gene
Fig. 2. The plasmid pBluescript. Indicated are characteristic sequences responsible for the
function of the vector (mcr = multiple cloning region).