STRUCTURAL AND CHEMICAL ARCHITECTURE OF HOST CELLS 49 



5. The Isolation and Composition of Chromosomes 



Starting with intact tissues or isolated nuclei, nuclear membranes have 

 been disrupted by vigorous mechanical treatment, such as that provided by 

 a Waring Blendor. The extent to which these treatments damage the internal 

 chromosomal structures or produce artifacts is not clear. Sonic vibrations 

 readily depolymerize DNA and the grinding of bacteria with alumina can 

 result in the complete solubilization of deoxyribonucleoprotein. It may be 

 noted that the shearing forces produced in a Waring Blendor readily convert 

 thymus DNA of average molecular weight 8 X 10® to a DNA of average 

 molecular weight 2 X 10® (Cohen, Hanlon, and Schachman, unpublished 

 observations). 



In any case, Claude and Potter (1943) and Mirsky and Pollister (1943) 

 showed that fine chromatin threads containing deoxyribonucleohistone may 

 be isolated from the disrupted resting nuclei of leukemic ceUs, erythrocytes, 

 and Hver. These threads contain over 90 % of the DNA of the cell. If these 

 threads are indeed the chromosomes of resting nuclei, this would be important 

 evidence for the theory of the continuity of chromosomes throughout the 

 life of the cell. 



Several workers have challenged the claim that these threads are condensed 

 chromosomes. Lamb (1950) states that various disruption techniques merely 

 draw out the nuclei into threads or fragments that bear Httle resemblance to 

 mitotic chromosomes. Schneider and Hogeboom (1951) report that the 

 addition of isotonic saline to Hver nuclei caused the precipitation of threads 

 within the previously homogeneous nuclei. If nuclei are isolated in sucrose 

 and disrupted, threads were not observed and over 60 % of the DNA was 

 not sedimentable at 60,000 g. 



However, Mirsky and Ris (1947, 1951) have examined their isolated 

 threads quite closely and consider that they have the visible characteristics 

 of chromosomes. Further, a number of individual types can be seen repeatedly. 

 Polli (1952) and Denues (1952, 1953) have supported this view. A close 

 similarity has been found between the threads from erythrocjrte nuclei and 

 the chromosomes observed in the metaphase plates of vertebrate cells. 

 Mitotic chromosomes also correspond closely in length with isolated chromo- 

 somes of the same species, which also show characteristic intraspecific 

 differences. 



Four types of compounds have been described in the "chromosomes." 

 Nucleohistone, consisting of DNA and histone, has been extracted at low 

 ionic strength, a process which is markedly facihtated by a prior treatment 

 of the chromosome with a chelating agent, such as citrate or ethylenediamine 

 tetraacetate (Mazia, 1954). The components of nucleohistone may be obtained 

 in the dissociated state by extraction in solutions of high ionic strength, 

 e.g., 1-2 M NaCl. The viscous solution produced in this way may be 



