Blunt End Ligation by T4 DNA Ligase, Duplex DNA, DNA Molecules, Palindrome, DNA Strands, Molecule Irrespective

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Chimeric DNA Molecular Probes and Gene Libraries Chimeric DNA Molecular Probes and Gene Libraries Introduction Restriction Enzymes for Cloning Technique of Restriction Mapping Restriction Clevage and Gel Electrophoresis Construction of a Restriction Map Use of Partial Digests end Labelling and Hybridization in Restriction Mapping Construction of Chimeric DNA Cloning of Plasmid Vectors Directional Cloning of DNA Fragments with Heterologus Protruding Termini Adding Poly dA at 3' Ends of DNA Clone Blunt End Ligation by T4 DNA Ligase Cloning in Phage λ and Cosmid Vectors Cloning in phage λ Vectors Cloning in Cosmid Vectors Hosts for Cloning Vectors Bacteria as Hosts Eukaryotes as Hosts Molecular Probes Preparation of Probes Genomic DNA Probes

Home >> Biotechnology and Genomics >> Chimeric DNA Molecular Probes and Gene Libraries >> Blunt End Ligation by T4 DNA Ligase

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Blunt end ligation by T4 DNA ligase

In this technique, a restriction enzyme is used to cut a duplex DNA at the same place in both the DNA strands. The broken ends are then used for joining with the two ends of another DNA molecule irrespective of the sequences present at the broken ends of the two DNA molecules. The T4 DNA ligase is used for this joining reaction. The disadvantage of this technique is that any two broken ends may join including those belonging to the same DNA molecule.

This leads to the production of a variety of products and one will have to select the desired product from a mixture of products. The blunt end ligation is .used for developing a method in which the DNA to be cloned can be easily retrieved whenever required.

Addition of a "linker" to a vector molecule

Addition to a Linker (carrying a Restriction site) to a Vector Molecule

This method makes use of short DNA duplexes (linkers), that contain EcoRI palindrome or some equivalent palindrome, which being small in size, can be synthesized chemically. These linkers can be linked to the blunt ends of vector DNA by blunt end ligation. This will allow the creation of an EcoRI site in the linker region of the vector, so that when a DNA segment is now cloned, the inserted DNA segment can be retrieved by cleavage with EcoRI.

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