XPC detect distortions = UvrA in E. coli

Formation of the bubble involves the helicase activities of the proteins XPA and XPD (the equivalent to UvrB in E. coli) and the single-strand-binding protein RPA. The bubble creates cleavage sites 5’ to the lesion for a nuclease known as ERCC1-XPF and 3’ to the lesion for nuclease XPG (representing the function of UvrC). In higher cells, the resulting DNA strand is 24–32 nucleotides long. As in bacteria, the DNA strand is released to create a gap that is filled in by the action of DNA polymerase and ligase.

XPB and XPD are both helicases; the XPB helicase is required for promoter melting during transcription, while the XPD helicase performs the unwinding function in NER

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#Unit 13. Methods in Biology

In ITP, the separation is carried out in a discontinuous buffer system. The ionized sample migrates between a leading electrolyte with a high mobility and a terminating – sometimes called trailing – ion with a low mobility, all of them migrating with the same speed.

#Id: 2475

#Unit 13. Methods in Biology

IEF takes place in a pH gradient and can only be used for amphoteric substances such as peptides and proteins. The charged molecules move toward the anode or the cathode until they reach a position in the pH gradient where their net charges are zero. This pH value is the “isoelectric point” (pI) of the substance.

#Id: 2476

#Unit 13. Methods in Biology

Agarose is one of the components of agar, which is a mixture of polysaccharides isolated from certain seaweeds and usually used at concentrations of between 1% and 3% in labs. 

#Id: 2477

#Unit 13. Methods in Biology

Agarose gels are formed  by suspending dry agarose in aqueous buffer, then boiling the mixture until a clear solution forms. 

#Id: 2478

#Unit 13. Methods in Biology

The most commonly used buffer for separating DNA is TRIS-acetate containing EDTA ( TAE) with a typical pH of 8.3 and used to dissolve the matrix (agarose) as well as the running buffer. 

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#Unit 13. Methods in Biology

Separation of RNA, TRIS-borate with EDTA ( TBE) is typically used due to superior buffering capacity (beneficial for long run times).