CsCl profiles

Figure 3.5. Aiialyiical CsCl density gradient profiles of four mouse DNA samples of different molecular weight. Molecular weights were measured from sedimentation velocity experiments except for the highest one which was estimated from the CsCl profile (From Macaya et at., 1976).

The second discovery was that isochores belonged to a small number of families characterized by distinct average base compositions and by other sequence features (see Table 3.4). The major DNA components were responsible for the discontinuities revealed by the CsCl analysis of CS2SO4 ligand fractions (Fig. 3.4) and derived from isochore families that caused discontinuities in the CsCl profiles of very high molecular weight samples (Fig. 3.5). 

Compositional patterns were called genome phenotypes (Bernard and Bernard , 1986a), because they are different in different vertebrate classes (see also Part 4). It should be stressed that, until the work of Thiery et al. (1976), the common belief was that, while DNA composition varied greatly among bacteria, animal species had a rather similar DNA composition (Kit, 1960). Although some small variations in modal buoyant densities and standard deviation of CsCl profiles had been detected in vertebrate DNAs, these could be due to different methylation levels (Kit, 1962). 

Figure 4.1 displays the analytical CsCl profiles of the fish DNAs studied by Bernard and Bernard (1990a). Only three DNA samples (4, 11, 12 all of them from Chondrichtyes) showed sizable amounts of resolved satellite bands, which were on the heavy, GC-rich side of the main band. Minor satellite bands were found in other DNA samples, mainly on the... 

The CsCl profiles of DNAs from amphibians (Fig. 4.7) show no satellite bands cryptic satellites may exist, however. For instance, in the case of X. laevis, the CsCl analysis of fractions from preparative centrifugation in Cs2S04/Ag showed (see Fig. 3.3) the presence of minor components banding at 1.705 and 1.706 g/cm and of a minute amount of a 1.712 g/cm component (Thiery et al., 1976). 

Figure 4.13. CsCl profiles of 18 rodent species (solid line) and their best fits by broadened exponential distributions (thin dashed lines). The abscissa shows modal buoyant density in g/cm the ordinate shows relative amounts of DNA. All profiles are normalized to unit area and shown in three columns in order of increasing modal buoyant density. No fits are shown for the two Geomyoidea profiles (see the original paper). The dashed vertical line, at 1.7015 g/cm3, indicates the mean buoyant density of the 18 species.

From Douady et al. (2000). Taxonomy of families and subfamilies follows Wilson and Reeder (1993, pp. 7-8). Sources are denoted by a. Douady et al. (2000) b, Sabeur et al. (1993) c, Mazrimas and Hatch (1972) d, Walker (1968) and Hennig and Walker (1970). Modal buoyant densities from sources c and d are approximate only and are shown in brackets, since they were estimated from published pictures of smoothed CsCl profiles means and other parameters could not be calculated from those profiles because of nonlinear responses (unreliable vertical scales). Two species with lower molecular weights are denoted by asterisks. 

The analysis of the localization of GC-poor DNA on metaphase and prometaphase chromosomes will be illustrated in some detail in order to provide an example of the procedure followed. Fig. 7.10 displays the CsCl profiles, the relative DNA level, and the proportion of the isochore families in the DNA fractions (Saccone et al., 1996) used as probes for the in situ hybridizations of GC-poor DNA. LI isochores are only present in the pellet DNA L2 isochores are distributed in the pellet and in fractions 1-3 fractions 4 and 5 contain almost exclusively DNA from FIl isochores. In the case of fraction 5 (as well as in the following fractions shown in Fig. 1 of Saccone et al., 1996), a light satellite peak corresponding to DNA from centromeric heterochromatin is present. The pellet DNA, essentially formed by LI isochores, hybridized on a subset of G bands (Fig. 7.11), which were called LI bands, and corresponded to the G1 and G2 bands, as we will name the two most intensely staining subsets of Francke s R bands (1994). In contrast, the pellet DNA is almost absent in the large majority of the FI3 4oo and H3 4oo bands. This is especially evident in chromosomes 15, 17, 19, and 22 (see Fig. 7.11). 

Figure 7. ), CsCl profiles of the human DNA fractions toiuaining the LI, L2 and HI isochore families (modified from Sacconc et ai 1996) fraction numbers (P stands for pellet), rcUitivcDNA content, modal buojant density and relative amounts of isochorc famitics are indicated. (From Federico et a1 2000).

Sixteen single-copy coding sequences were hybridized on DNA fractions from shallow CsCl gradients. GC levels of the fractions corresponding to hybridization peaks are indicated on the analytical CsCl profile in Fig. 9.2A. The GC and GC3 levels of the coding sequences are shown in Fig. 9.2B. They cover most of the compositional spectrum of the... 

Fig. 9.3A shows a plot of GC3 levels of the sixteen coding sequences analyzed against the GC levels of the corresponding long sequences (GC fractions) in which the coding sequences were localized. The regression equation was used to position the GC3 distribution of the coding sequences available in data bank (about 100 sequences) relative to the CsCl profile (Fig. 9.3B). 

Figure 9.6. The GC histogram of large genomic sequences (> 50 kb) from Drosophila (A) and human (B) are superimposed on the CsCl profiles of the corresponding DNAs, as obtained by analytical ultracentrifugation (from Thiery et al., 1976, and Zoubak et ah, 1996, respectively). Buoyant densities were converted to GC levels according to Schildkraut et al. (1962). (From Jabbari and Bernardi, 2000).

Indeed, it was shown in Fig. 4.9 and pointed out in Part 4, Chapter 3, that within the suborder of snakes (which was taken into consideration because it comprises the largest number of analyzed subfamilies or genera see Tables 4A and 4B), the range of asymmetry of CsCl profiles is much wider than those of entire classes of vertebrates, fishes/amphibians or mammals/birds. The lack of similarity in asymmetry (and compositional patterns) in the snakes is a strong indication that compositional patterns developed independently in small taxons (subfamilies, genera). Our suggestion is that this independent development took place under the effect of different body temperatures. Several observations lend support to this explanation 1) different small taxa of reptiles are charaeterized by different body... 

Figure R4(6.4). Distribution of integrated IITLV-I and IIBV viral sequences in the human genotne. The CsCl profile ofhuman liSA is shown. Arrows indicate the GC level of viral sequences, expressed sequences are in red, non-expressed sequences in yellow.

Douady C., Cards N., Clay O., Catzeflis F. and Bernardi G. (2000). Diversity and phylogenetic implications of CsCl profiles from rodent DNAs. Mol. Phylogen. Evol. 17 219-230. 

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