cas1, cas2, and cas4 required to create protospacer sequences Cas1 is  integrase, whereas Cas2 is a ribonuclease. In contrast, of other Cas proteins, Cas6 is involved in expression and processing of the CRISPR cluster and Cas3 in the interference of viral infection.

#Id: 2779

#Unit 13. Methods in Biology

General structure and component of a chromatographic system

A stationary phase – typically an aqueous buffer with sufficient ionic strength to maintain a soluble protein.

A column -

An injector system: To deliver test samples to the top of the column in a reproducible manner.

A detector with data acquisition - To give a continuous record of the presence of the analytes in the eluate as it emerges from the column.

A fraction collector: For collecting the separated analytes for further biochemical studies.

#Id: 2780

#Unit 13. Methods in Biology

The basis of all forms of chromatography is the distribution or partition coefficient (Kd), which describes the way in which a compound (the analyte) distributes between two immiscible phases.

#Id: 2781

#Unit 13. Methods in Biology

A chromatogram is a pictorial record of the detector response as a function of elution volume or retention time, It consists of a series of peaks or bands.

#Id: 2782

#Unit 13. Methods in Biology

The retention time tR for each analyte has two components

 i - dead time, tM. &

ii - adjusted retention time, t’R.

#Id: 2783

#Unit 13. Methods in Biology

t’R is characteristic of the analyte and is the difference between the observed retention time and the dead time:

#Id: 2784

#Unit 13. Methods in Biology

Retention factor (k) is simply the additional time that the analyte takes to elute from the column relative to an unretained or excluded analyte that does not interact with the stationary phase and which, by definition, has a k value of 0. Thus