06-18-2014, 03:37 AM
Vectors Based on Bacteriophage M13
Phage M13 has some advantages but also some disadvantages concerning the usage for genetic experiments and based on them, it should be chosen only in some cases.
The first good thing about M13 is that its RF (replicative form) is similar to plasmid, so it can be isolated and manipulated using the same techniques. Moreover, single-stranded DNA of M13, which is produced during the infection, is useful in techniques like DNA sequencing by dideoxy method. Sequencing is very important in genetic experiments, and this feature of M13 bacteriophage makes it a good target for a potential vector.
The difference between phage lambda and M13 is in their genome. M13 does not have any non-essential genes making this one of its greatest downsides. Moreover, its genome is filled up very effectively, so its intergenic region (available for manipulation) is only 507 base pairs long. However, its genome is already pretty small (6407 base pairs).
Intergenic region in M13 is used to insert a polylinker/lacZ sequence into the vectors based on M13, which enables the X-gal screening system (blue white selection) making detection of recombinants easier. When M13 is grown on a bacterial lawn, plaques appear because of the reduction in growth of host cells, which can be picked up for further analysis.
Another disadvantage of vectors based on bacteriophage M13 is the amount of exogenous genetic material they can accept. Even though the capsid of M13 is actually determined by the size of its genome, it still cannot accept large fragments of exogenous DNA. In fact, anything longer than 1.5 kb is considered to be too much for M13 based vectors because it makes them loose their cloning efficiency. This problem is bypassed simply by using M13 vectors for cloning and sequencing of small DNA fragments; especially in the situations where the ease of purification is needed, or when potential sequencing is planned later on.
Phage M13 has some advantages but also some disadvantages concerning the usage for genetic experiments and based on them, it should be chosen only in some cases.
The first good thing about M13 is that its RF (replicative form) is similar to plasmid, so it can be isolated and manipulated using the same techniques. Moreover, single-stranded DNA of M13, which is produced during the infection, is useful in techniques like DNA sequencing by dideoxy method. Sequencing is very important in genetic experiments, and this feature of M13 bacteriophage makes it a good target for a potential vector.
The difference between phage lambda and M13 is in their genome. M13 does not have any non-essential genes making this one of its greatest downsides. Moreover, its genome is filled up very effectively, so its intergenic region (available for manipulation) is only 507 base pairs long. However, its genome is already pretty small (6407 base pairs).
Intergenic region in M13 is used to insert a polylinker/lacZ sequence into the vectors based on M13, which enables the X-gal screening system (blue white selection) making detection of recombinants easier. When M13 is grown on a bacterial lawn, plaques appear because of the reduction in growth of host cells, which can be picked up for further analysis.
Another disadvantage of vectors based on bacteriophage M13 is the amount of exogenous genetic material they can accept. Even though the capsid of M13 is actually determined by the size of its genome, it still cannot accept large fragments of exogenous DNA. In fact, anything longer than 1.5 kb is considered to be too much for M13 based vectors because it makes them loose their cloning efficiency. This problem is bypassed simply by using M13 vectors for cloning and sequencing of small DNA fragments; especially in the situations where the ease of purification is needed, or when potential sequencing is planned later on.
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