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Homopolymer tailing is a method in which homopolymer deoxynucleotides tails are added to blunt-ended polymers in order to join a double-stranded DNA fragment to a cloning vehicle. There will be an explanation of TA cloning, along with the TA sub-cloning method. The process is done by avoiding the use of restriction enzymes. At the end of this guide, analysis about ta brown gene cloning and TA cloning vector could be assessed.
Concept of TA Cloning
Discussion of TA Cloning Method
The TA Cloning method is done with the help of terminal transferase activity of some specific DNA polymerases such as Taq polymerase. In this process, the insert is produced in the form of a PCR. TA Cloning can be considered as one of the simplest and most efficient methods of cloning and subcloning of PCR products. This method of cloning is simple and can be considered as one of the most efficient methods of cloning, and includes Taq polymerase.
Explanation of TA Sub-cloning Methods along with TA Cloning
The TA sub-cloning method can be easily modified hence the T-vector that is used in one cloning method can be used in other processes. This is done in order to clone double-stranded DNA fragments. In this case, the PCR products are amplified by double-stranded DNA polymerase, as well as blunt and sticky-ended DNA fragments are included. Therefore, it can be summarised that the TA cloning method is a similar convenient and labour-saving sub-cloning method. On the other hand, in case of homopolymer tailing, blunt ended DNA fragments are ligated with the help of enzymes such as terminal deoxynucleotidyl transferase. In this process, a nucleotide is repeatedly added to the 3 Prime ends of double-stranded DNA molecules. Therefore, this process enables the TA cloning to perform easily and effectively.
Discussion of the TA Brown Gene Cloning
The TA cloning procedure begins by placing the insert in a PCR reaction using taq polymer and adds a single A onto the end of the PCR product. After this, the vector of the PCR mixes with the complementary 3’deoxythymidine (T) overhang. Then the DNA ligase is added to connect to the vector and insert. The same TA cloning vector can be used to clone any segment of the PCR amplified DNA. It does not require purifying or cutting the complementary vector as it is done for the restriction enzyme cloning.
According to TA Brown, Gene cloning is a phenomenon in which a fragment of DNA that contains the gene is cloned. After that, it is inserted into a circular DNA molecule that is called a vector. After completion of the mentioned process, a Recombinant DNA molecule is produced through gene cloning. The gene is transported to a host cell with the help of the vector and usually, the TA vector is a bacterium. In addition to that, different types of living cells can also be used as vectors in this process. At the time when the vector is inserted into the host cell, it gets multiplied and produces a number of identical copies. The copies that it produces not only include the parent cells but also give the genes that it carries. At the time when the host cell is divided, copies of the Recombinant DNA molecules are passed to progeny, and application of the vector takes place even further. As a result, a large number of cell divisions or a clone is produced.
TA Cloning Vector
TA vectors used in cloning are linearized plasmids that are treated in order to help PCR products by adding T-overhangs to the A overhang. TA cloning vectors can be used in order to clone any segment of DNA that is amplified by a PCR. The use of T-vector ensures that the cloning process is direct and highly efficient because it has single 3 Prime T overhangs on both ends. This method of sub-cloning of DNA fragments is useful, especially at the time when compatible restriction sites are unavailable. In this process, DNA fragments are cloned from one vector to another for getting a successful clone. Directional cloning is also possible in this method with the help of hemi-phosphorylation of both the T-vectors as well as the inserts. At the time when a single T vector is at a hand, any DNA fragments can be cloned with a lot more efficiency.
Conclusion
As per the above analysis, it can be concluded that TA cloning can be considered as one of the most effective sub-cloning processes that are also easier to use by the researchers. In the gene, the cloning process replication of a DNA fragment happens and that is one of the most important parts of gene cloning with the help of vectors. Recombinant DNA molecules take a major part in this process and a large number of cell divisions take place resulting in DNA cloning.
