Cloning vector

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The pGEX-3x plasmid is a popular cloning vector.

A cloning vector is a small DNA vehicle that carries a foreign DNA fragment. The insertion of the fragment into the cloning vector is done by treating the vehicle and the foreign DNA with the same restriction enzyme, then ligating the fragments together. There are many types of cloning vectors. Plasmids and bacteriophages (such as phage λ) are perhaps most commonly used for this purpose. Other types of cloning vectors include bacterial artificial chromosomes (BACs) and yeast artificial chromosomes (YACs).

Common Features

Most commercial cloning vectors have a couple of key features that have made their use in molecular biology so widespread.

Usually, the main purpose of cloning vector is the controlled expression of a particular gene inside a convient host organism (eg. E. coli). Control of expression can be very important, it is usually desirable to insert the target DNA in to a site that is under the control of a particular promoter. Some often used promoters are T7 promoters, lac promoters (bla promoter) and cauliflower mosaic virus's 35s promoter (for plant vectors). To allow for convient and favorable insertions, most cloning vectors have had nearly all their restriction sites engineered out of them and a multiple cloning site(MCS) inserted that contains many restriction sites. MCSs allow for insertions of DNA into the vector to be targeted and possibly directed in a chosen orientatiton. A selectable marker, such as antibiotic resistance [eg. beta-lactamase (see figure)] is often included in the vector to identify postivly transformed cells. All inserted DNA (plasmids etc.) need an origin of replication (ori; not shown in figure); high strigency ori's preferably for cloning vectors.

Other possible features of cloning vectors maybe, vir genes for plant transformation, intergrase sites for chromosomal insertion, lac Z alpha fragment for blue-white selection, and/or in-frame genes attached to the MCS for recombinant proteins [eg. Green fluorescent protein (GFP) or glutathione S-transferase (see figure)].



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