Cucurbit Genetics Cooperative Report 10:4-6 (article 3) 1987
Composition of Nuclear DNA in Cucumis species
College of Horticulture, Kerala Agricultural University, Vellanikkara 680651,
Department of Agricultural Botany, University College of Wales, Aberystwyth,
The genus Cucumis comprises about thirty species of Asian and
African origin. The diploid species are dibasic with haploid chromosome
numbers, n=7 or n=12. About 17 percent of Cucumis species are polyploids
and their chromosome numbers vary from 4x=48 to 6x=72. The 2C DNA amounts
among diploid species range narrowly between 1.37 pg and 2.48 pg, and polyploids
between 2.48 pg and 3.38 pg. The quantitative DNA changes associated with
speciation in Cucumis has affected all chromosomes within each chromosome
The nuclear DNA composition of four Cucumis species viz., C. melo, C. metuliferus, C. anguria and C. sativus were studied by buoyant density gradient analysis and thermal
denaturation analysis. The method for DNA extraction was similar to that
described by Bendich and Anderson (1). Analytical CsCl density gradient
analysis was done in a Centriscan - 75 ultracentrifuge. Purified DNA (1.5
mg from each Cucumis species) was centrifuged to equilibrium in neutral
CsCl (density 1.710 g/ml). Micrococcus lysodeikticus DNA of known
buoyant density was included as a marker. For thermal denaturation analysis,
DNA samples were melted in 0.1 x SSC (SSC = 0.15M NaCl + 0.015M trisodium
citrate, pH 7.0) in a fully automated SP 1800 Spectrophotometer equipped
with an electronically heated cell block. The increase in absorbance (hyperchromicity)
was recorded automatically in a digital printout for every 0.25°C
increase in temperature. The absorbance measurements after correction for
solvent expansion (4) were plotted as a ratio, At /A25 (absorbance at temperature 't' divided by initial absorbance at 25°C)
The buoyant density profiles for all species were asymmetric (Figure
1). The asymmetry suggests the presence of satellite DNA sequences with
different average buoyant density. Satellite DNA sequences have been identified
and characterized previously in the genomes of C. sativus and C. melo (1,2,3). The buoyant densities for the total main
band DNA ranged from 1.6909 g/ml for C. anguria to 1.6920
g/ml for C. sativus, C. melo and C. metuliferus have intermediate values (Table 1). The melting profiles when compared under
strictly identical ionic conditions gave us information about the base ratio
of DNA as well as about heterogeneity, if any, in the dispersion of base
pairs. Tm (the temperature corresponding to half the final increase
in relative absorbance), base ratio (G+C content) and base compositional
heterogeneity estimated from the melting profile are also given in Table
1. Tm values ranged between 69.5°C and 70.25°C.
Guanine + Cytosine content (G+C) of DNA estimated from Tm was
similar to the values derived from buoyant density (7) and ranged from 38.06
to 39.89%. C. sativus DNA has an entirely different estimate
of base compositional heterogeneity (14.67%) compared with the other three Cucumis species. Detailed results of the isolation, characterization
and in situ hybridization of satellite DNA sequences from Cucumis species will be published elsewhere (6).
Table 1. Nuclear DNA composition of Cucumis species
Figure 1. Buoyant density profiles for nuclear DNA of Cucumis species with
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