page_title: Multiple cloning site
section_title: Creating a multiple cloning site
hierarchical_section_title: Multiple cloning site / Creating a multiple cloning site
caption_attribution_description: English: This image shows the process of putting a multiple cloning site into a plasmid vector.
context_page_description: A multiple cloning site, also called a polylinker, is a short segment of DNA which contains many restriction sites - a standard feature of engineered plasmids. Restriction sites within an MCS are typically unique, occurring only once within a given plasmid. The purpose of a MCS in a plasmid is to allow a piece of DNA to be inserted into that region. A MCS is found in a variety of vectors, including cloning vectors to increase the number of copies of target DNA, and in expression vectors to create a protein product. In expression vectors, the MCS is located downstream of the promoter.
context_section_description: In some instances, a vector may not contain a MCS. Rather, a MCS can be added to a vector. The first step is designing complementary oligonucleotide sequences that contain restriction enzyme sites along with additional bases on the end that are complementary to the vector after digesting. Then the oligonucleotide sequences can be annealed and ligated into the digested and purified vector. The digested vector is cut with a restriction enzyme that complements the oligonucleotide insert overhangs. After ligation, transform the vector into bacteria and verify the insert by sequencing. This method can also be used to add new restriction sites to a multiple cloning site.
caption: A diagram showing the process of inserting a multiple cloning site into a plasmid vector.
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