Adding Poly dA at 3' Ends of DNA Clone, Enzyme Terminal Transferase, DNA Sequence Cloned, DNA Molecular, Poly Dc Regenerate Recognition

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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 >> Adding Poly dA at 3' Ends of DNA Clone

Your Ad Here	Adding poly dA at 3' ends of the vector and poly dT at the 3' ends of DNA clone In this method, DNA can be cut at the desired position both in the vector and in the clone, without staggered cleavage. Using precursor dA TP, poly dA is added at both the 3' cut ends of the vector with the help of enzyme terminal transferase. Cloning of DNA segment in a plasmid vectror by poly dA; poly dT tailing technique
Similarly using precursor dTTP poly dT is added at both the 3' cut ends of the DNA sequence to be cloned. The vector and clone can then be joined by annealing the poly dA with poly dT tails and then ligating them using DNA ligase enzyme In this techinque, there is no chance of reannealing between the two cut ends of the same DNA molecule or of the two similar DNA molecules, so that one of the disadvantages of the first method has been overcome in this case.
However, in this technique it will not be easy to retrieve the cloned DNA, because the recognition site of the enzyme has been lost in this case due to insertion of poly dA and poly dT. But if poly dG and poly dC are used instead (which serve the same purpose), cloned sequence can be retrieved, since poly dG and poly dC regenerate the recognition site for the enzyme PstI..	Your Ad Here

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