Transposon as tools for genetic engineering

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Genomics Summer Semester 2011        Evaluation Essay        Dmytro Dvornikov

Genomics Summer Semester 2011.

Evaluation essay

Transposons have been widely used as genetic tools to study many model systems. However only recently have they been used in mammals. Why? Discuss the recent applications of transposition to mammalian systems and consider the advantages and disadvantages of transposition compared to other genetic methods.

Abstract

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For decades transposon-based mutagenesis and transgenesis were used in invertebrate model organisms such as D. melanogaster and C. elegans. This approach offers numerous advantages over conventional methods of viral transduction or non-viral techniques. However, up until recently such methods were not available for vertebrates. Only after an extinct transposable element Sleeping Beauty (SB) was recovered from fish genome in 1997 situation has changed drastically. SB proved to be efficient in somatic, germinal and embryonic stem cells of different vertebrate model animals. Sleeping Beauty was found to be an indispensable tool for loss-of-function and cancer screens, generation of virus-free induced pluripotent stem cells (iPSCs) and human gene therapy. Further development of SB and other transposon systems in vertebrates will help to fill up the gap existing in this field between vertebrate and invertebrate model organisms.

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Transposable Elements

        Transposons are pieces of DNA that can move themselves within genomes. Most of them have a relatively simple structure: a gene encoding the transposase protein is flanked by terminal inverted repeats. The transposase can be easily replaced with any sequence of interest. Such transposon can only move if transposase is trans supplemented.

        There are two main types of transposons. First is retrotransposons. They utilize replicative "copy-and-paste" mechanism: donor element is transcribed, new copy is converted back to DNA and inserts somewhere in the genome. The most common members of that group are LINEs and SINEs. The second group is called DNA transposons. They lack RNA stage in their life cycle. DNA transposons use "cut-and-paste" mechanism to jump through the genome.

Transposons in comparison with other genetic methods

        DNA transposons proved to be quite useful as genetic tools for easy and controllable way of genome manipulation. They have a distinct set of advantages over other methods of genome modification.

        Before transposon era two main ways of generation transgenic organisms existed: DNA microinjection and viral transduction. However each of them have their own drawbacks. Thus, DNA microinjection suffers from low rates of genomic integration. Furthermore, injected DNA tends to integrate as a concatemer that results in transgene silencing [1]. Viral vectors, in turn, was found to be mutagenic, since they tend to integrate into genes or induce rearrangements and deletions [2]. Whereas non-integrating adeno-associating viral vectors require repeating rounds of administration that often cause severe immune response [3].

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        In contrast to viruses, transposons are not infectious and therefore can propagate only within the host cell. Thus, transposons evolved to cause as less harm as possible. They have low preference for insertion into genes and usually fairly low transpositional activity. Additionally, transposition is effective from conventional plasmids and therefore do not cause any immunological complications [4]. All that make transposons safe, effective tools for genome manipulations that are easy to use and engineer.

        However, there are certain basic requirements for any transposon as a genetic tool in any particular model organism. First of all, it is a sufficient level ...

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