Search results
Results from the WOW.Com Content Network
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.
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. The most often used are genes pIII or pVIII of bacteriophage M13. [5]
The T7 promoter region allows large-scale in vitro T7 transcription to transcribe the DNA library into an mRNA library, which provides templates for the in vitro translation reaction later. The ribosomal binding site in the 5’-untranslated region (5’ UTR) is designed according to the in vitro translation system to be used.
The 'helper' phage infects the bacterial host by first attaching to the host cell's pilus and then, after attachment, transporting the phage genome into the cytoplasm of the host cell. Inside the cell, the phage genome triggers production of single stranded phagemid DNA in the cytoplasm. This phagemid DNA is then packaged into phage particles.
Phage typing is based on the specific binding of phages to antigens and receptors on the surface of bacteria and the resulting bacterial lysis or lack thereof. [4] The binding process is known as adsorption. [5] Once a phage adsorbs to the surface of a bacteria, it may undergo either the lytic cycle or the lysogenic cycle. [6]
Structure of phage ΦX174 capsid Schematic drawing of a Sinsheimervirus (aka Phix174microvirus) virion. The phi X 174 (or ΦX174) bacteriophage is a single-stranded DNA virus that infects Escherichia coli.
M13 is one of the Ff phages (fd and f1 are others), a member of the family filamentous bacteriophage ().Ff phages are composed of circular single-stranded DNA (), which in the case of the m13 phage is 6407 nucleotides long and is encapsulated in approximately 2700 copies of the major coat protein p8, and capped with about 5 copies each of four different minor coat proteins (p3 and p6 at one ...
Phage-assisted continuous evolution (PACE) is a phage-based technique for the automated directed evolution of proteins. It relies on relating the desired activity of a target protein with the fitness of an infectious bacteriophage which carries the protein's corresponding gene.