Volume 3

Gene expression / Benjamin Lewin.

  • Lewin, Benjamin.
Date:
1974-1977
Catalogue details

Licence: Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)

Credit: Gene expression / Benjamin Lewin. Source: Wellcome Collection.

    62/952 (page 42)
    formants and the concentration of DNA (up to the plateau level beyond which the number of transformants does not increase). Double transfor mants may arise in either of two ways. For unlinked markers, two DNA molecules each carrying a different donor genetic marker may interact with a single cell; this cell may then become transformed for both charac teristics, due to two independent integration events. In this case, the kinetics of double transformation display two hit kinetics; and when transformation frequencies are expressed relative to the number of com petent cells (that is, rather than to the total number of viable cells), the frequency of double transformation is the product of the two single transformation frequencies. By a reversal of this argument, in fact the more usual sequence in which the calculation is performed, it is possible to calculate the proportion of competent cells by comparing the frequency of the double transformation event with the product of the single trans formation frequencies (now expressed relative to the total cell count). For linked markers, both may be carried on a single molecule of DNA, in which case the recipient may be transformed for both characteristics in a single event. This results, of course, in an increased frequency of double transformation, dependent upon single hit kinetics. As we have just implied, linkage between two markers cannot be assumed because the frequency of double transformation is greater than the product of the relevant single transformation frequencies; for spurious linkage will result whenever the proportion of competent cells is less than 100%. As Goodgal and Herriot (1961) pointed out, comparison of double with single transformation frequencies can be valid only when expressed relative to the number of competent cells. Thus for two markers trans formed independently (assuming that both donor markers are present in the transforming DNA preparation in the same concentration, as will be the case whenever they were originally located on one genome, but too far apart to be linked, or if two different DNA preparations are used in equal amounts): N a „ N b _ N ab f f f so that N ah = — X N„ or f = N a X N t N ab