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  Section: General Biotechnology / Genes & Genetic Engineering
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Techniques of Genetic Engineering


Animal viruses
A variety of animal viruses are known, for example, simian virus 40 (SV40), adinovirus, retrovirus (ssRNA), vaccinia virus, etc. They can increase the efficiency of animal cell transformation if they are used as vector. A special feature of viruses is that they contain the strong promoters which can possibly bring about expression of inserted DNA fragment of the viruses. The most commonly used virus is SV40 which contains a circular DNA of about 5.2 Kb. In addition to ori region, the DNA contains early genes and late genes. Early genes are required for DNA replication, whereas late genes encode coat protein of the virus.

Like phage λ, the amount of DNA to be packed into virus capsid is limited. The genome has 3 identified non-essential regions; hence the use of non-defective viral vectors is limited. The defective virus vectors have missing genes whose infection process has to be helped by viral DNA constituting functional copies of missing genes. If the cos cells (the host monkey cells) are used as host, they can be transformed by a plasmid carrying the early region of SV40 with a defective replication origin, cos cells express wild type large T antigen and can support the growth of SV40. Recombinants containing foreign DNA inserted into early region can be propagated without the need of helper virus (Glover, 1984). In this way, the recombinant and helper can be separately recovered from the host.



Gene cloning in prokaryotes


Isolation of DNA to be cloned 


Insertion of DNA fragment into vector 



Use of restriction Linkers



Use of homopolymer tails


Transfer of recombinant DNA into bacterial cells


Selection of clones



Colony hybridization techniques



In vitro translation technique



Immunological tests



Blotting Techniques


Recovery of cells


Expression of cloned DNA



Shine-Dalgano sequence



Expression vectors

Gene cloning in eukaryotes


Plant cells






Filamentous fungi



Agrobacterium plasmids



Plant cell transformation



Plant cell transformation by ultrasonication



Liposome mediated gene transfer


Animal cell  



Animal viruses






Particle bombardment






Direct transformation

Site directed mutagenesis


Methods of mutagenesis


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