Other Vector Systems
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Vector Systems
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Vectors |
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Biology |
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Vectors |
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As we have become more sophisticated in our ability to manipulate and characterize DNA sequences, new vector systems have been developed for specific uses. This is a short look at some of these other systems.
A major thrust of vector development has been to create vectors that will handle larger foreign DNA inserts, aiming to reduce the number of recombinants it is necessary to look at in order to identify a specific DNA sequence. Cosmid vectors were among the first large insert cloning vehicles developed.
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A cosmid is a plasmid that contains a cos site from the lambda genome.
The vector replicates as a plasmid Ligation of cosmid vector and foreign DNA fragments (SauIIIA partial digest fragments 45 kb in size) is similar to ligation into a lambda substitution vector.
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This concatemer is then packaged into viral particles (remember packaging is cos site to cos site) and these are used to infect E. coli where the cosmid vector replicates using the ColE1 orignin of replication. Phage packaging serves only to select for recombinant molecules and to transfer these long DNA molecues (50 kb total) into the bacterial host (50 kb fragments transform very inefficiently while phage infection is very efficient).
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Very Large Prokaryotic Vector Systems
Recently, the need for prokaryotic vectors capable of replicating very large foreign DNA fragments A few years ago, a Bacterial Artificial Chromosome (BAC) vector was developed allowing foreign DNA fragements over 1 MB to be propagated in E. coli. Standard methods for transforming E coli rely on the chemical preparation of the E coli to put them in a 'transformation competent' state. Getting very large molecules across the cell membrane relies on an alternative mechanism called electroporation. This involves mixing DNA with E coli cells and then exposing the mixture to a high-voltage pulse. The high voltage induces massive membrane rearrangements and the coincident uptake of DNA which is independant of size. BACs have been an important tool for the human genome sequencing project.
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Eukaryotic Vector Systems
Numerous vector systems have been developed for use in eukaryotic systems. Yeast Vectors Mammalian Vectors Finally, artificial chromosome vectors have been developed for use in yeast (YACs) and are being developed for use in mammalian cells (MACs).
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