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Phage display cycle. 1) fusion proteins for a viral coat protein + the gene to be evolved (typically an antibody fragment) are expressed in bacteriophage. 2) the library of phage are washed over an immobilised target. 3) the remaining high-affinity binders are used to infect bacteria. 4) the genes encoding the high-affinity binders are isolated.
Biopanning involves 4 major steps for peptide selection. [4] The first step is to have phage display libraries prepared. This involves inserting foreign desired gene segments into a region of the bacteriophage genome, so that the peptide product will be displayed on the surface of the bacteriophage virion.
Cosmids are plasmids that incorporate a segment of bacteriophage λ DNA that has the cohesive end site (cos) which contains elements required for packaging DNA into λ particles. Under apt origin of replication (ori), it can replicate as a plasmid.
They contain a cosN site where DNA is nicked at each strand, 12 bp apart, by terminase. This causes linearization of the circular cosmid with two "cohesive" or "sticky ends" of 12bp. (The DNA must be linear to fit into a phage head.) The cosB site holds the terminase while it is
The first step in Gateway cloning is the preparation of a Gateway Entry clone. There are a few different ways to make entry clone. Gateway attB1 and attB2 sequences are added to the 5' and 3' end of a gene fragment, respectively, using gene-specific PCR primers and PCR amplification. The PCR amplification products are then mixed with a propriet
The integration of phage λ takes place at a special attachment site in the bacterial and phage genomes, called att λ. The sequence of the bacterial att site is called attB, between the gal and bio operons, and consists of the parts B-O-B', whereas the complementary sequence in the circular phage genome is called attP and consists of the parts ...
In 1938, Craigie and Yen adapted Vi phages by selective propagation and used them at their critical test dilutions to differentiate 11 types of B. typhosus. [19] In 1943, Felix and Callow extended the method to Salmonella paratyphi B . in 1943 and differentiated 12 types with 11 phages. [ 20 ]
Molecular cloning takes advantage of the fact that the chemical structure of DNA is fundamentally the same in all living organisms. Therefore, if any segment of DNA from any organism is inserted into a DNA segment containing the molecular sequences required for DNA replication, and the resulting recombinant DNA is introduced into the organism from which the replication sequences were obtained ...