Inserted DNA sequences

These vectors integrate stably into the host s DNA, and they are not known to cause disease in humans. Their disadvantage is that they accept only a relatively small inserted DNA sequence (about 5 kb, as opposed to 8 kb in a retrovirus and 30 kb in an adenovirus). 

Return to the plasmid window, place cursor at the cloning site, and paste the insertion DNA sequence from the clipboard (i.e., Edit -> Paste). 

INOC = [(4-pyridinylmethoxy)carbonyl)], 241 myo-Inositol selective glycosylation, 270 Inserted DNA sequences, 243-245 Insulin synth. by genetic engineering, 244-245 Insulin -chain chemical synthesis, 239-240 Introns, 242... 

PCR can also be used to modify DNA sequences using primers differing at one or several positions from the target sequence. This is possible because PCR does not require perfect complementarity of a primer to the sequence flanking the target. Since all of the PCR products contain the primer sequence, an insertion or deletion can thus be incorporated into the product by modifying a primer. It is also possible to add new sequences to the 5 -ends of the primers. Modified or additional genetic information may thus be multiplied and transr ported. 

DNA polymerase enzymes all synthesize DNA by adding deoxynucleotides to the free 3 -OH group of an RNA or DNA primer sequence. The identity of the inserted nucleotide is deterrnined by its abiHty to base-pair with the template nucleic acid. The dependence of synthesis on a primer oligonucleotide means that synthesis of DNA proceeds only in a 5%o V direction if only one primer is available, all newly synthesized DNA sequences begin at the same point. 

FIGURE 13.2 Foreign DNA sequences can be inserted into plasmid vectors by opening the circnlar plasmid with a restriction endonnclease. The ends of the linearized plasmid DNA are then joined with the ends of a foreign sequence, reclosing the circle to create a chimeric plasmid. 

There is a normal variation of DNA sequence just as is true of more obvious aspects of human structute. Variations of DNA sequence, polymorphisms, occur approximately once in evety 500 nucleotides, or about 10 times per genome. There are without doubt deletions and insertions of DNA as well as single-base substitutions. In healthy people, these alterations obviously occur in noncoding regions of DNA or at sites that cause no change in function of the encoded protein. This heritable polymorphism of DNA structure can be associated with certain diseases within a large kindred and can be used to search for the specific gene involved, as is illustrated below. It can also be used in a variety of applications in forensic medicine. 

Cosmid A plasmid into which the DNA sequences from bacteriophage lambda that are necessary for the packaging of DNA (cos sites) have been inserted this permits the plasmid DNA to be packaged in vitro. 

More extensive changes in the DNA sequence (often referred to as macrolesions) can also occur. Deletions result in the loss of part of the DNA sequence. Insertions add extra base pairs to a gene. Transversions occur when a segment ofthe DNA is reversed and duplications occur when a segment ofthe DNA is repeated. Some ofthese changes also result in frameshifts. 

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