Mutagenesis by PCR using SOE reaction
SOE stands for Splicing by overhang extensions ,is an immensely useful tools in molecular biology today for creating point mutations in gene of interest and also to make chimeric proteins (fusion proteins).The important point of SOE is designing of primer sequences used for amplifying two or more DNA fragments to be fused.The primers are designed in a way that resultant PCR products have complimentary ends so that they can act as primers for each other in subsequent reaction.Then when you mix the resultant pcr products and run a pcr reaction again using primers at ends of two DNA fragments (5′ and 3′),results in fusion of two fragments of PCR into one double stranded DNA molecule.This can be compared to cloning with used of ligations and restriction endonucleases.
SOE reactions are widely used to create point mutations and producing chimeric proteins.For making point mutations,deletions or insertions one need to design the altered primers with mutation to be introduced in their sequence.As the primers will have overhangs they can be joined in next round of pcr reaction.The resultant will be a full length DNA fragment with mutation of interest.
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